What's hot today:
Thursday, June 30th, 2022 - Adult Development
What's hot today
|Alba, V., Carthew, J. E., Carthew, R. W. and Mani, M. (2021). Global constraints within the developmental program of the Drosophila wing. Elife 10. PubMed ID: 34180394
Organismal development is a complex process, involving a vast number of molecular constituents interacting on multiple spatio-temporal scales in the formation of intricate body structures. Despite this complexity, development is remarkably reproducible and displays tolerance to both genetic and environmental perturbations. This robustness implies the existence of hidden simplicities in developmental programs. Using the Drosophila wing as a model system, a new quantitative strategy was developed that enables a robust description of biologically salient phenotypic variation. Analyzing natural phenotypic variation across a highly outbred population and variation generated by weak perturbations in genetic and environmental conditions, a highly constrained set of wing phenotypes was observed. Remarkably, the phenotypic variants can be described by a single integrated mode that corresponds to a non-intuitive combination of structural variations across the wing. This work demonstrates the presence of constraints that funnel environmental inputs and genetic variation into phenotypes stretched along a single axis in morphological space. These results provide quantitative insights into the nature of robustness in complex forms while yet accommodating the potential for evolutionary variations. Methodologically, a general strategy for finding such invariances in other developmental contexts is introduced.
|Allen, A. M. and Sokolowski, M. B. (2021). Expression of the foraging gene in adult Drosophila melanogaster. J Neurogenet: 1-21. PubMed ID: 34382904
The foraging gene in Drosophila melanogaster, which encodes a cGMP-dependent protein kinase, is a highly conserved, complex gene with multiple pleiotropic behavioral and physiological functions in both the larval and adult fly. Adult foraging expression is less well characterized than in the larva. This study characterized foraging expression in the brain, gastric system, and reproductive systems using a T2A-Gal4 gene-trap allele. In the brain, fforaging expression appears to be restricted to multiple sub-types of glia. This glial-specific cellular localization of foraging was supported by single-cell transcriptomic atlases of the adult brain. foraging is extensively expressed in most cell types in the gastric and reproductive systems. Multiple cis-regulatory elements responsible for parts of the observed expression patterns were identified by a nested cloned promoter-Gal4 analysis. The mapped cis-regulatory elements were consistently modular when comparing the larval and adult expression patterns. These new data using the T2A-Gal4 gene-trap and cloned foraging promoter fusion GAL4's are discussed with respect to previous work using an anti-FOR antibody, which is shown here to be non-specific. Future studies of foraging's function will consider roles for glial subtypes and peripheral tissues (gastric and reproductive systems) in foraging's pleiotropic behavioral and physiological effects.
|Wyatt, B. H., Amin, N. M., Bagley, K., Wcisel, D., Dush, M. K., Yoder, J. A. and Nascone-Yoder, N. M. (2021). Single-minded 2 is required for left-right asymmetric stomach morphogenesis. Development. PubMed ID: 34180499
The morphogenesis of left-right (LR) asymmetry is a critical phase of organogenesis. In the digestive tract, the development of anatomical asymmetry is first evident in the leftward curvature of the stomach. To elucidate the molecular events that shape this archetypal laterality, transcriptome analyses was performed of the left versus right sides of the developing stomach in frog embryos. Besides the known LR gene pitx2, the only gene found to be expressed asymmetrically throughout all stages of curvature was single-minded2 (sim2), a Down Syndrome-related transcription factor and homolog of a Drosophila gene (sim) required for LR asymmetric looping of the fly gut. sim2 functions downstream of LR patterning cues to regulate key cellular properties and behaviors in the left stomach epithelium that drive asymmetric curvature. These results reveal unexpected convergent cooption of single-minded genes during the evolution of LR asymmetric morphogenesis, and have implications for dose-dependent roles of laterality factors in non-laterality-related birth defects.
|Pan, X. and O'Connor, M. B. (2021). Coordination among multiple receptor tyrosine kinase signals controls Drosophila developmental timing and body size. Cell Rep 36(9): 109644. PubMed ID: 34469735
In holometabolous insects, metamorphic timing and body size are controlled by a neuroendocrine axis composed of the ecdysone-producing prothoracic gland (PG) and its presynaptic neurons (PGNs) producing PTTH. Although PTTH/Torso signaling is considered the primary mediator of metamorphic timing, recent studies indicate that other unidentified PGN-derived factors also affect timing. This study demonstrates that the receptor tyrosine kinases anaplastic lymphoma kinase (Alk) and PDGF and VEGF receptor-related (Pvr), function in coordination with PTTH/Torso signaling to regulate pupariation timing and body size. Both Alk and Pvr trigger Ras/Erk signaling in the PG to upregulate expression of ecdysone biosynthetic enzymes, while Alk also suppresses autophagy by activating phosphatidylinositol 3-kinase (PI3K)/Akt. The Alk ligand Jelly belly (Jeb) is produced by the PGNs and serves as a second PGN-derived tropic factor, while Pvr activation mainly relies on autocrine signaling by PG-derived Pvf2 and Pvf3. These findings illustrate that a combination of juxtacrine and autocrine signaling regulates metamorphic timing, the defining event of holometabolous development.
|Yang, D. W., Mok, J. W., Telerman, S. B., Amson, R., Telerman, A. and Choi, K. W. (2021). Topoisomerase II is regulated by translationally controlled tumor protein for cell survival during organ growth in Drosophila. Cell Death Dis 12(9): 811. PubMed ID: 34453033
Regulation of cell survival is critical for organ development. Translationally controlled tumor protein (TCTP) is a conserved protein family implicated in the control of cell survival during normal development and tumorigenesis. Previously,a human Topoisomerase II (TOP2) was identified as a TCTP partner, but its role in vivo has been unknown. To determine the significance of this interaction, their roles in developing Drosophila organs was examined. Top2 RNAi in the wing disc leads to tissue reduction and caspase activation, indicating the essential role of Top2 for cell survival. Top2 RNAi in the eye disc also causes loss of eye and head tissues. Tctp RNAi enhances the phenotypes of Top2 RNAi. The depletion of Tctp reduces Top2 levels in the wing disc and vice versa. Wing size is reduced by Top2 overexpression, implying that proper regulation of Top2 level is important for normal organ development. The wing phenotype of Tctp RNAi is partially suppressed by Top2 overexpression. This study suggests that mutual regulation of Tctp and Top2 protein levels is critical for cell survival during organ development.
|Pojer, J. M., Manning, S. A., Kroeger, B., Kondo, S. and Harvey, K. F. (2021). The Hippo pathway uses different machinery to control cell fate and organ size. iScience 24(8): 102830. PubMed ID: 34355153
The Hippo pathway is a conserved signaling network that regulates organ growth and cell fate. One such cell fate decision is that of R8 photoreceptor cells in the Drosophila eye, where Hippo specifies whether cells sense blue or green light. This study shows that only a subset of proteins that control organ growth via the Hippo pathway also regulate R8 cell fate choice, including the STRIPAK complex, Tao, Pez, and 14-3-3 proteins. Furthermore, key Hippo pathway proteins were primarily cytoplasmic in R8 cells rather than localized to specific membrane domains, as in cells of growing epithelial organs. Additionally, Warts was the only Hippo pathway protein to be differentially expressed between R8 subtypes, while central Hippo pathway proteins were expressed at dramatically lower levels in adult and pupal eyes than in growing larval eyes. Therefore, this study has revealed several important differences in Hippo signaling in the contexts of organ growth and cell fate.
Wednesday September 29th - Signaling
|McParland, A., Moulton, J., Brann, C., Hale, C., Otis, Y. and Ganter, G. (2021). The brinker repressor system regulates injury-induced nociceptive sensitization in Drosophila melanogaster. Mol Pain 17: 17448069211037401. PubMed ID: 34399634
Chronic pain is a debilitating condition affecting millions of people worldwide, and an improved understanding of the pathophysiology of chronic pain is urgently needed. Nociceptors are the sensory neurons that alert the nervous system to potentially harmful stimuli such as mechanical pressure or noxious thermal temperature. When an injury occurs, the nociceptive threshold for pain is reduced and an increased pain signal is produced. This process is called nociceptive sensitization. This sensitization normally subsides after the injury is healed. However, dysregulation can occur which results in sensitization that persists after the injury has healed. This process is thought to perpetuate chronic pain. The Hedgehog (Hh) signaling pathway has been previously implicated in nociceptive sensitization in response to injury in Drosophila melanogaster. Downstream of Hh signaling, the Bone Morphogenetic Protein (BMP) pathway has also been shown to be necessary for this process. This study describes a role for nuclear components of BMP's signaling pathway in the formation of injury-induced nociceptive sensitization. Brinker (Brk), and Schnurri (Shn) were suppressed in nociceptors using an RNA-interference (RNAi) "knockdown" approach. Knockdown of Brk resulted in hypersensitivity in the absence of injury, indicating that it normally acts to suppress nociceptive sensitivity. Animals in which transcriptional activator Shn was knocked down in nociceptors failed to develop normal allodynia after ultraviolet irradiation injury, indicating that Shn normally acts to promote hypersensitivity after injury. These results indicate that Brk-related transcription regulators play a crucial role in the formation of nociceptive sensitization and may therefore represent valuable new targets for pain-relieving medications.
|Nichols, R., Pittala, K., Leander, M., Maynard, B., Nikolaou, P. and Marciniak, P. (2021). The
myosuppressin structure-activity relationship for cardiac contractility
and its receptor interactions support the presence of a ligand-directed
signaling pathway in heart. Peptides: 170641. PubMed ID: 34453985
The structural conservation and activity of the myosuppressin cardioinhibitory peptide across species suggests it plays an important role in physiology, yet much remains unknown regarding its signaling. Drosophila melanogaster myosuppressin (dromyosuppressin, DMS; TDVDHVFLRF-NH(2)) decreases cardiac contractility through a G protein-coupled receptor, DMS-R2. This study showed the DMS N-terminus amino acids influence its structure-activity relationship (SAR), yet how they act is not established. It was predicted that myosuppressin N-terminal amino acids played a role in activity and signaling. This hypothesis was tested in the beetle, Zophobas atratus, using a semi-isolated heart bioassay to explore SAR in a different Order and focus on cardiac signaling. A series of myosuppressin truncated analogs was generated by removing the N-terminal residue and measuring the activity of each structure on cardiac contractility. While DVDHVFLRF-NH(2) decreased cardiac contractility, it was found VDHVFLRF-NH(2), DHVFLRF-NH(2), and HVFLRF-NH(2) increased activity. In contrast, VFLRF- NH(2) decreased activity and FLRF-NH(2) was inactive. Next, molecular docking data was analyzed, and it was found the active truncated analogs interacted with the 3-6 lock in DMS-R2, the myosuppressin cardiac receptor, disrupting the salt bridge between H114 and E369, and K289 and Q372. Further, the docking results showed the inhibitory effect on contractility may be associated with contact to Y78, while the analogs that increased contractility lacked this interaction. The data from this study demonstrated N-terminal amino acids played a role in myosuppressin activity and signaling suggesting the cardiac receptor can be targeted by biased agonists. These myosuppressin cardiac contractility data and predicted receptor interactions describe the presence of functional selectivity in a ligand-directed signaling pathway in heart.
|Wang, W., Lin, H., Zheng, E., Hou, Z., Liu, Y., Huang, W., Chen, D., Feng, J., Li, J. and Li, L. (2021). Regulation of survivin protein stability by USP35 is evolutionarily conserved. Biochem Biophys Res Commun 574: 48-55. PubMed ID: 34438346
Survivin is the key component of the chromosomal passenger complex and plays important roles in the regulation of cell division. Survivin has also been implicated in the regulation of apoptosis and tumorigenesis. Although the survivin protein has been reported to be degraded by a ubiquitin/proteasome-dependent mechanism, whether there is a DUB that is involved in the regulation of its protein stability is largely unknown. Using an expression library containing 68 deubiquitinating enzymes, this study found that ubiquitin-specific-processing protease 35 (USP35) regulates survivin protein stability in an enzymatic activity-dependent manner. USP35 interacted with and promoted the deubiquitination of the survivin protein. USP38, an ortholog of USP35 encoded by the human genome, is also able to regulate survivin protein stability. Moreover, this study found that the deubiquitinating enzyme DUBAI, the Drosophila homolog of human USP35, is able to regulate the protein stability of Deterin, the Drosophila homolog of survivin. Interestingly, USP35 also regulated the protein stability of Aurora B and Borealin which are also the component of the chromosomal passenger complex. By regulating protein stabilities of chromosomal passenger complex components, USP35 regulates cancer cell proliferation. Taken together, this work uncovered an evolutionarily conserved relationship between USP35 and survivin that might play an important role in cell proliferation.
|Beatty, J. S., Molnar, C., Luque, C. M., de Celis, J. F. and Martin-Bermudo, M. D. (2021). EGFRAP encodes a new negative regulator of the EGFR acting in both normal and oncogenic EGFR/Ras-driven tissue morphogenesis. PLoS Genet 17(8): e1009738. PubMed ID: 34411095
Activation of Ras signaling occurs in ~30% of human cancers. However, activated Ras alone is insufficient to produce malignancy. Thus, it is imperative to identify those genes cooperating with activated Ras in driving tumoral growth. This work identified a novel EGFR inhibitor, which was named EGFRAP, for EGFR adaptor protein. Elimination of EGFRAP potentiates activated Ras-induced overgrowth in the Drosophila wing imaginal disc. EGFRAP interacts physically with the phosphorylated form of EGFR via its SH2 domain. EGFRAP is expressed at high levels in regions of maximal EGFR/Ras pathway activity, such as at the presumptive wing margin. In addition, EGFRAP expression is up-regulated in conditions of oncogenic EGFR/Ras activation. Normal and oncogenic EGFR/Ras-mediated upregulation of EGRAP levels depend on the Notch pathway. Elimination of EGFRAP does not affect overall organogenesis or viability. However, simultaneous downregulation of EGFRAP and its ortholog PVRAP results in defects associated with increased EGFR function. Based on these results, it is proposed that EGFRAP is a new negative regulator of the EGFR/Ras pathway, which, while being required redundantly for normal morphogenesis, behaves as an important modulator of EGFR/Ras-driven tissue hyperplasia. It is suggested that the ability of EGFRAP to functionally inhibit the EGFR pathway in oncogenic cells results from the activation of a feedback loop leading to increase EGFRAP expression. This could act as a surveillance mechanism to prevent excessive EGFR activity and uncontrolled cell growth.
|Viswanathan, R., Hartmann, J., Pallares Cartes, C. and De Renzis, S. (2021). Desensitisation of Notch signalling through dynamic adaptation in the nucleus. Embo J: e107245. PubMed ID: 34396565
During embryonic development, signalling pathways orchestrate organogenesis by controlling tissue-specific gene expression programmes and differentiation. Although the molecular components of many common developmental signalling systems are known, the current understanding of how signalling inputs are translated into gene expression outputs in real-time is limited. This study employed optogenetics to control the activation of Notch signalling during Drosophila embryogenesis with minute accuracy and follow target gene expression by quantitative live imaging. Light-induced nuclear translocation of the Notch Intracellular Domain (NICD) causes a rapid activation of target mRNA expression. However, target gene transcription gradually decays over time despite continuous photo-activation and nuclear NICD accumulation, indicating dynamic adaptation to the signalling input. Using mathematical modelling and molecular perturbations, this study showed that this adaptive transcriptional response fits to known motifs capable of generating near-perfect adaptation and can be best explained by state-dependent inactivation at the target cis-regulatory region. Taken together, these results reveal dynamic nuclear adaptation as a novel mechanism controlling Notch signalling output during tissue differentiation.
|Wang, R., Xie,
H., Yang, L., Wang, P., Chen, M. M., Wu, H. Y., Liao, Y. L., Wang, M.
Y., Wang, Q., Gong, X. X., Cheng, Q., Cheng, L., Xie, F. Y., Bi, C. L.
and Fang, M. (2021). Naked cuticle inhibits wingless signaling in Drosophila wing development. Biochem Biophys Res Commun 576: 1-6. PubMed ID: 34474244
Wnt signaling is one of the major signaling pathways that regulate cell differentiation, tissue patterning and stem cell homeostasis and its dysfunction causes many human diseases, such as cancer. It is of tremendous interests to understand how Wnt signaling is regulated in a precise manner both temporally and spatially. Naked cuticle (Nkd) acts as a negative-feedback inhibitor for Wingless (Wg, a fly Wnt) signaling in Drosophila embryonic development. However, the role of Nkd remains controversial in later fly development, particularly on the canonical Wg pathway. This study shows that nkd is essential for wing pattern formation, such that both gain and loss of nkd result in the disruption of Wg target expression in larvae stage and abnormal adult wing morphologies. Furthermore, it was demonstrated that a thirty amino acid fragment in Nkd, identified previously in Wharton lab, is critical for the canonical Wg signaling, but is dispensable for Wg/planar cell polarity pathway. Putting aside the pleiotropic nature of nkd function, i.e. its role in the Decapentaplegic signaling, it is concluded that Nkd universally inhibits the canonical Wg pathway across a life span of Drosophila development.
Tuesday, September 28th - Adult Physiology
|Tuthill, B. F., Quaglia, C. J., O'Hara, E. and Musselman, L. P. (2021). Loss of Stearoyl-CoA Desaturase 1 leads to cardiac dysfunction and lipotoxicity. J Exp Biol. PubMed ID: 34423827
Diets high in carbohydrates are associated with type 2 diabetes and its comorbidities, including hyperglycemia, hyperlipidemia, obesity, hepatic steatosis and cardiovascular disease. This study used a high-sugar diet to study the pathophysiology of diet-induced metabolic disease in Drosophila melanogaster. High-sugar diets produce hyperglycemia, obesity, insulin resistance, and cardiomyopathy in flies along with ectopic accumulation of toxic lipids, or lipotoxicity. Stearoyl-CoA desaturase 1 is an enzyme that contributes to long-chain fatty acid metabolism by introducing a double bond into the acyl chain. Knockdown of stearoyl-CoA desaturase 1 in the fat body reduced lipogenesis and exacerbated pathophysiology in flies reared on high-sugar diets. These flies exhibited dyslipidemia and growth deficiency in addition to defects in cardiac and gut function. The lipidome of these flies was assessed using tandem mass spectrometry to provide insight into the relationship between potentially lipotoxic species and type 2 diabetes-like pathophysiology. Oleic acid supplementation is able to rescue a variety of phenotypes produced by stearoyl-CoA desaturase 1 RNAi, including fly weight, triglyceride storage, gut development, and cardiac failure. Taken together, these data suggest a protective role for monounsaturated fatty acids in diet-induced metabolic disease phenotypes.
|Leech, T., McDowall, L., Hopkins, K. P., Sait, S. M., Harrison, X. A. and Bretman, A. (2021). Social
environment drives sex and age-specific variation in Drosophila
melanogaster microbiome composition and predicted function. Mol Ecol. PubMed ID: 34494339
The composition of the microbiome (the assemblage of symbiotic microorganisms within a host) is determined by environmental factors and the host's immune, endocrine and neural systems. The social environment could alter host microbiomes extrinsically by affecting transmission between individuals. Alternatively, intrinsic effects arising from interactions between the microbiome and host physiology (the microbiota-gut-brain axis) could translate social stress into dysbiotic microbiomes, with consequences for host health. This study investigated how manipulating social environments during larval and adult life-stages altered the microbiome composition of Drosophila melanogaster fruit flies. Social contexts that particularly alter the development and lifespan of males were used, predicting that any intrinsic social effects on the microbiome would therefore be sex-specific. The presence of adult males during the larval stage significantly altered the microbiome of pupae of both sexes. In adults, same-sex grouping increased bacterial diversity in both sexes. Importantly, the microbiome community structure of males was more sensitive to social contact at older ages, an effect partially mitigated by housing focal males with young rather than coaged groups. Functional analyses suggest that these microbiome changes impact ageing and immune responses. This is consistent with the hypothesis that the substantial effects of the social environment on individual health are mediated through intrinsic effects on the microbiome, and provides a model for understanding the mechanistic basis of the microbiota-gut-brain axis.
|Lukacs, A., Thomae,
A. W., Krueger, P., Schauer, T., Venkatasubramani, A. V., Kochanova, N.
Y., Aftab, W., Choudhury, R., Forne, I. and Imhof, A. (2021). The Integrity of the HMR complex is necessary for centromeric binding and reproductive isolation in Drosophila. PLoS Genet 17(8): e1009744. PubMed ID: 34424906
Postzygotic isolation by genomic conflict is a major cause for the formation of species. Despite its importance, the molecular mechanisms that result in the lethality of interspecies hybrids are still largely unclear. The genus Drosophila, which contains over 1600 different species, is one of the best characterized model systems to study these questions. It has been shown that the expression levels of the two hybrid incompatibility factors Hmr and Lhr diverged in the two closely related Drosophila species, D. melanogaster and D. simulans, resulting in an increased level of both proteins in interspecies hybrids. The overexpression of the two proteins also leads to mitotic defects, a misregulation in the expression of transposable elements and decreased fertility in pure species. This work describes a distinct six subunit protein complex containing HMR and LHR and analyse the effect of Hmr mutations on complex integrity and function. These experiments suggest that HMR needs to bring together components of centromeric and pericentromeric chromatin to fulfil its physiological function and to cause hybrid male lethality.
|Kam, J. H., Hogg, C., Fosbury, R., Shinhmar, H. and Jeffery, G. (2021). Mitochondria
are specifically vulnerable to 420nm light in Drosophila which
undermines their function and is associated with reduced fly mobility. PLoS One 16(9): e0257149. PubMed ID: 34478469
Increased blue light exposure has become a matter of concern as it has a range of detrimental effects, but the mechanisms remain unclear. Mitochondria absorb short wavelength light but have a specific absorbance at 420nm at the lower end of the human visual range. This 420nm absorption is probably due to the presence of porphyrin. This study examined the impact of 420nm exposure on Drosophila melanogaster mitochondria and its impact on fly mobility. Daily 15 mins exposures for a week significantly reduced mitochondrial complex activities and increased mitochondrial inner membrane permeability, which is a key metric of mitochondrial health. Adenosine triphosphate (ATP) levels were not significantly reduced and mobility was unchanged. There are multiple options for energy/time exposure combinations, but single 420nm exposure of 3h was applied to increase the probability of an effect on ATP and mobility, and both were significantly reduced. ATP and mitochondrial membrane permeability recovered and over corrected at 72h post exposure. However, despite this, normal mobility did not return. Hence, the effect of short wavelengths on mitochondrial function is to reduce complex activity and increasing membrane permeability, but light exposure to reduce ATP and to translate into reduced mobility needs to be sustained.
|Heys, C., Fisher, A. M., Dewhurst, A. D., Lewis, Z. and Lize, A. (2021). Exposure to foreign gut microbiota can facilitate rapid dietary shifts. Sci Rep 11(1): 16791. PubMed ID: 34408232
Dietary niche is fundamental for determining species ecology; thus, a detailed understanding of what drives variation in dietary niche is vital for predicting ecological shifts and could have implications for species management. Gut microbiota can be important for determining an organism's dietary preference, and therefore which food resources they are likely to exploit. Evidence for whether the composition of the gut microbiota is plastic in response to changes in diet is mixed. Also, the extent to which dietary preference can be changed following colonisation by new gut microbiota from different species is unknown. This study used Drosophila spp. to show that: (1) the composition of an individual's gut microbiota can change in response to dietary changes, and (2) ingestion of foreign gut microbes can cause individuals to be attracted to food types they previously had a strong aversion to. Thus, this study exposes a mechanism for facilitating rapid shifts in dietary niche over short evolutionary timescales.
|Koyama, T., Terhzaz, S., Naseem, M. T., Nagy, S., Rewitz, K., Dow, J. A. T., Davies, S. A. and Halberg, K. V. (2021). A nutrient-responsive hormonal circuit mediates an inter-tissue program regulating metabolic homeostasis in adult Drosophila. Nat Commun 12(1): 5178. PubMed ID: 34462441
Animals maintain metabolic homeostasis by modulating the activity of specialized organs that adjust internal metabolism to external conditions. However, the hormonal signals coordinating these functions are incompletely characterized. This study shows that six neurosecretory cells in the Drosophila central nervous system respond to circulating nutrient levels by releasing Capa hormones, homologs of mammalian neuromedin U, which activate the Capa receptor (CapaR) in peripheral tissues to control energy homeostasis. Loss of Capa/CapaR signaling causes intestinal hypomotility and impaired nutrient absorption, which gradually deplete internal nutrient stores and reduce organismal lifespan. Conversely, increased Capa/CapaR activity increases fluid and waste excretion. Furthermore, Capa/CapaR inhibits the release of glucagon-like adipokinetic hormone from the corpora cardiaca, which restricts energy mobilization from adipose tissue to avoid harmful hyperglycemia. These results suggest that the Capa/CapaR circuit occupies a central node in a homeostatic program that facilitates the digestion and absorption of nutrients and regulates systemic energy balance.
Monday, September 27th - Embryonic Development
|Yu, J. C., Balaghi, N., Erdemci-Tandogan, G., Castle, V. and Fernandez-Gonzalez, R. (2021). Myosin cables control the timing of tissue internalization in the Drosophila embryo. Cells Dev: 203721. PubMed ID: 34271226
Compartment boundaries prevent cell mixing during animal development. In the Drosophila embryo, the mesectoderm is a group of glial precursors that separate ectoderm and mesoderm, forming the ventral midline. Mesectoderm cells undergo one round of oriented divisions during axis elongation and are eventually internalized approximately 6 h later. Using spinning disk confocal microscopy and image analysis, this study found that after dividing, mesectoderm cells reversed their planar polarity. The polarity factor Bazooka was redistributed to mesectoderm-mesectoderm cell interfaces, and the molecular motor non-muscle Myosin II and its upstream activator Rho-kinase (Rok) accumulated at mesectoderm-ectoderm (ME) interfaces, forming supracellular cables flanking the mesectoderm on either side of the tissue. Laser ablation revealed the presence of increased tension at ME cables, where Myosin was stabilized, as shown by fluorescence recovery after photobleaching. Laser nanosurgery was used to reduce tension at the ME boundary, and Myosin fluorescence decreased rapidly, suggesting a role for tension in ME boundary maintenance. Mathematical modelling predicted that increased tension at the ME boundary was necessary to prevent the premature establishment of contacts between the two ectodermal sheets on opposite sides of the mesectoderm, thus controlling the timing of mesectoderm internalization. The model was validated in vivo: Myosin inhibition disrupted the linearity of the ME boundary and resulted in early internalization of the mesectoderm. These results suggest that the redistribution of Rok polarizes Myosin and Bazooka within the mesectoderm to establish tissue boundaries, and that ME boundaries control the timely internalization of the mesectoderm as embryos develop.
|Shlemov, A., Alexandrov, T., Golyandina, N., Holloway, D., Baumgartner, S. and Spirov, A. V. (2021). Quantification reveals early dynamics in Drosophila maternal gradients. PLoS One 16(8): e0244701. PubMed ID: 34411119
The Bicoid (Bcd) protein is a primary determinant of early anterior-posterior (AP) axis specification in Drosophila embryogenesis. This study produced confocal microscope images of whole early embryos, stained for bcd mRNA or the Staufen (Stau) protein involved in its transport. Each profile was quantified by a two- (or three-) exponential equation. The parameters of these equations were used to analyze the early developmental dynamics of bcd. Analysis of 1D profiles was compared with 2D intensity surfaces from the same images. This approach reveals strong early changes in bcd and Stau, which appear to be coordinated. Three stages in early development can be unambiguously discriminated using the exponential parameters: pre-blastoderm (1-9 cleavage cycle, cc), syncytial blastoderm (10-13 cc) and cellularization (from 14A cc). Key features which differ in this period are how fast the first exponential (anterior component) of the apical profile drops with distance and whether it is higher or lower than the basal first exponential. Both bcd and Stau show several redistributions in the head cytoplasm, quite probably related to nuclear activity. The continued spreading of bcd can be tracked from the time of nuclear layer formation (later pre-blastoderm) to the later syncytial blastoderm stages by the progressive loss of steepness of the apical anterior exponential (for both bcd and Stau). Finally, at the beginning of cc14 (cellularization stage) a distinctive flip is seen from the basal anterior gradient being higher to the apical gradient being higher (for both bcd and Stau). Quantitative analysis reveals substantial (and correlated) bcd and Stau redistributions during early development, supporting that the distribution and dynamics of bcd mRNA are key factors in the formation and maintenance of the Bcd protein morphogenetic gradient.
|Bhide, S., Gombalova, D., Monke, G., Stegmaier, J., Zinchenko, V., Kreshuk, A., Belmonte, J. M. and Leptin, M. (2021). Mechanical competition alters the cellular interpretation of an endogenous genetic program. J Cell Biol 220(11). PubMed ID: 34449835
The intrinsic genetic program of a cell is not sufficient to explain all of the cell's activities. External mechanical stimuli are increasingly recognized as determinants of cell behavior. In the epithelial folding event that constitutes the beginning of gastrulation in Drosophila, the genetic program of the future mesoderm leads to the establishment of a contractile actomyosin network that triggers apical constriction of cells and thereby tissue folding. However, some cells do not constrict but instead stretch, even though they share the same genetic program as their constricting neighbors. This study shows that tissue-wide interactions force these cells to expand even when an otherwise sufficient amount of apical, active actomyosin is present. Models based on contractile forces and linear stress-strain responses do not reproduce experimental observations, but simulations in which cells behave as ductile materials with nonlinear mechanical properties do. These models show that this behavior is a general emergent property of actomyosin networks in a supracellular context, in accordance with experimental observations of actin reorganization within stretching cells.
|Cai, X., Rondeel, I. and Baumgartner, S. (2021). Modulating the bicoid gradient in space and time. Hereditas 158(1): 29. PubMed ID: 34404481
The formation of the Bicoid (Bcd) gradient in the early Drosophila is one of the most fascinating observations in biology and serves as a paradigm for gradient formation, yet its mechanism is still not fully understood. Two distinct models were proposed in the past, the SDD and the ARTS model. This study defines novel cis- and trans-acting factors that are indispensable for gradient formation. The first one is the poly A tail length of the bcd mRNA where this study demonstrates that it changes not only in time, but also in space. Posterior bcd mRNAs were shown to possess a longer poly A tail than anterior ones and this elongation is likely mediated by wispy (wisp), a poly A polymerase. Consequently, modulating the activity of Wisp results in changes of the Bcd gradient, in controlling downstream targets such as the gap and pair-rule genes, and also in influencing the cuticular pattern. Attempts to modulate the Bcd gradient by subjecting the egg to an extra nuclear cycle, i.e. a 15(th) nuclear cycle by means of the maternal haploid (mh) mutation showed no effect, neither on the appearance of the gradient nor on the control of downstream target. This suggests that the segmental anlagen are determined during the first 14 nuclear cycles. Finally, the Cyclin B (CycB) gene was identified as a trans-acting factor that modulates the movement of Bcd such that Bcd movement is allowed to move through the interior of the egg. This analysis demonstrates that Bcd gradient formation is far more complex than previously thought requiring a revision of the models of how the gradient is formed.
Andersen, P., Sulistio, E., Liu, X., Murphy, S., Kannan, S., Nam, L.,
Miyamoto, W., Tampakakis, E., Hibino, N., Uosaki, H. and Kwon, C. (2021). Noncanonical Notch signals have opposing roles during cardiac development. Biochem Biophys Res Commun 577: 12-16. PubMed ID: 34487959
The Notch pathway is an ancient intercellular signaling system with crucial roles in numerous cell-fate decision processes across species. While the canonical pathway is activated by ligand-induced cleavage and nuclear localization of membrane-bound Notch, Notch can also exert its activity in a ligand/transcription-independent fashion, which is conserved in Drosophila, Xenopus, and mammals. However, the noncanonical role remains poorly understood in in vivo processes. This study shows that increased levels of the Notch intracellular domain (NICD) in the early mesoderm inhibit heart development, potentially through impaired induction of the second heart field (SHF), independently of the transcriptional effector RBP-J. Similarly, inhibiting Notch cleavage, shown to increase noncanonical Notch activity, suppressed SHF induction in embryonic stem cell (ESC)-derived mesodermal cells. In contrast, NICD overexpression in late cardiac progenitor cells lacking RBP-J resulted in an increase in heart size. This study suggests that noncanonical Notch signaling has stage-specific roles during cardiac development.
|Song, Y. and Hyeon, C. (2021). Cost-precision trade-off relation determines the optimal morphogen gradient for accurate biological pattern formation. Elife 10. PubMed ID: 34402427
Spatial boundaries formed during animal development originate from the pre-patterning of tissues by signaling molecules, called morphogens. The accuracy of boundary location is limited by the fluctuations of morphogen concentration that thresholds the expression level of target gene. Producing more morphogen molecules, which gives rise to smaller relative fluctuations, would better serve to shape more precise target boundaries; however, it incurs more thermodynamic cost. In the classical diffusion-depletion model of morphogen profile formation, the morphogen molecules synthesized from a local source display an exponentially decaying concentration profile with a characteristic length λ. It is hypothesized that in order to attain a precise profile with the minimal cost, λ should be roughly half the distance to the target boundary position from the source. Remarkably, it was found that the profiles of Bcd, Wg, Hh, and Dpp, morphogens that pattern the Drosophila embryo and wing imaginal disk, are formed with nearly optimal λ. This finding underscores the thermodynamic cost as a key physical constraint in the morphogen profile formation in Drosophila development.
Friday, September 24th - Cytoskeleton and Junctions
|Schroeder, C. M., Tomlin, S. A., Mejia Natividad, I., Valenzuela, J. R., Young, J. M. and Malik, H. S. (2021). An actin-related protein that is most highly expressed in Drosophila testes is critical for embryonic development. Elife 10. PubMed ID: 34282725
Most actin-related proteins (Arps) are highly conserved and carry out well-defined cellular functions in eukaryotes. However, many lineages like Drosophila and mammals encode divergent non-canonical Arps whose roles remain unknown. To elucidate the function of non-canonical Arps, this study focused on Arp53D, which is highly expressed in testes and retained throughout Drosophila evolution. Arp53D localizes to fusomes and actin cones, two germline-specific actin structures critical for sperm maturation, via a unique N-terminal tail. Surprisingly, it was found that male fertility is not impaired upon Arp53D loss, yet population cage experiments reveal that Arp53D is required for optimal fitness in Drosophila melanogaster. To reconcile these findings, focus was placed on Arp53D function in ovaries and embryos where it is only weakly expressed. Under heat stress Arp53D-knockout (KO) females lay embryos with reduced nuclear integrity and lower viability; these defects are further exacerbated in Arp53D-KO embryos. Thus, despite its relatively recent evolution and primarily testis-specific expression, non-canonical Arp53D is required for optimal embryonic development in Drosophila.
|Denk-Lobnig, M., Totz, J. F., Heer, N. C., Dunkel, J. and Martin, A. C. (2021). Combinatorial patterns of graded RhoA activation and uniform F-actin depletion promote tissue curvature. Development 148(11). PubMed ID: 34124762
During development, gene expression regulates cell mechanics and shape to sculpt tissues. Epithelial folding proceeds through distinct cell shape changes that occur simultaneously in different regions of a tissue. Using quantitative imaging in Drosophila melanogaster, this study investigate how patterned cell shape changes promote tissue bending during early embryogenesis. The transcription factors Twist and Snail combinatorially regulate a multicellular pattern of lateral F-actin density that differs from the previously described Myosin-2 gradient. This F-actin pattern correlates with whether cells apically constrict, stretch or maintain their shape. Tthe Myosin-2 gradient and F-actin depletion do not depend on force transmission, suggesting that transcriptional activity is required to create these patterns. The Myosin-2 gradient width results from a gradient in RhoA activation that is refined through the balance between RhoGEF2 and the RhoGAP C-GAP. These experimental results and simulations of a 3D elastic shell model show that tuning gradient width regulates tissue curvature.
L., Jagtap, P. K. A., Cyrklaff, A., Vaishali, Lapouge, K., Sehr, P.,
Perez, K., Heber, S., Low, C., Hennig, J. and Ephrussi, A. (2021). Molecular basis of mRNA transport by a kinesin-1-atypical tropomyosin complex. Genes Dev 35(13-14): 976-991. PubMed ID: 34140355
Kinesin-1 carries cargos including proteins, RNAs, vesicles, and pathogens over long distances within cells. The mechanochemical cycle of kinesins is well described, but how they establish cargo specificity is not fully understood. Transport of oskar mRNA to the posterior pole of the Drosophila oocyte is mediated by Drosophila kinesin-1, also called kinesin heavy chain (Khc), and a putative cargo adaptor, the atypical tropomyosin, aTm1. How the proteins cooperate in mRNA transport is unknown. This study presents the high-resolution crystal structure of a Khc-aTm1 complex. The proteins form a tripartite coiled coil comprising two in-register Khc chains and one aTm1 chain, in antiparallel orientation. aTm1 binds to an evolutionarily conserved cargo binding site on Khc, and mutational analysis confirms the importance of this interaction for mRNA transport in vivo. Furthermore, this study demonstrates that Khc binds RNA directly and that it does so via its alternative cargo binding domain, which forms a positively charged joint surface with aTm1, as well as through its adjacent auxiliary microtubule binding domain. Finally, aTm1 was shown to plays a stabilizing role in the interaction of Khc with RNA, which distinguishes aTm1 from classical motor adaptors.
|Sharma, M., Jiang, T., Jiang, Z. C., Moguel-Lehmer, C. E. and Harris, T. J. (2021). Emergence of a smooth interface from growth of a dendritic network against a mechanosensitive contractile material. Elife 10. PubMed ID: 34423780
Structures and machines require smoothening of raw materials. Self-organized smoothening guides cell and tissue morphogenesis and is relevant to advanced manufacturing. Across the syncytial Drosophila embryo surface, smooth interfaces form between expanding Arp2/3-based actin caps and surrounding actomyosin networks, demarcating the circumferences of nascent dome-like compartments used for pseudocleavage. This study found that forming a smooth and circular boundary of the surrounding actomyosin domain requires Arp2/3 in vivo. To dissect the physical basis of this requirement, the interacting networks were reconstituted using node-based models. In simulations of actomyosin networks with local clearances in place of Arp2/3 domains, rough boundaries persisted when myosin contractility was low. With addition of expanding Arp2/3 network domains, myosin domain boundaries failed to smoothen, but accumulated myosin nodes and tension. After incorporating actomyosin mechanosensitivity, Arp2/3 network growth locally induced a surrounding contractile actomyosin ring that smoothened the interface between the cytoskeletal domains, an effect also evident in vivo. In this way, a smooth structure can emerge from the lateral interaction of irregular active materials.
|Falo-Sanjuan, J. and Bray, S. J. (2021). Membrane architecture and adherens junctions contribute to strong Notch pathway activation. Development. PubMed ID: 34486648
The Notch pathway mediates cell-to-cell communication in a variety of tissues, developmental stages and organisms. Pathway activation relies on the interaction between transmembrane ligands and receptors on adjacent cells. As such, pathway activity could be influenced by the size, composition or dynamics of contacts between membranes. The initiation of Notch signalling in the Drosophila embryo occurs during cellularization, when lateral cell membranes and adherens junctions are first being deposited, allowing the investigation of the importance of membrane architecture and specific junctional domains for signaling. By measuring Notch dependent transcription in live embryos it was established that Notch initiates while lateral membranes are growing and that signalling onset correlates with a specific phase in their formation. However, the length of the lateral membranes per se was not limiting. Rather, the adherens junction likely play an important role in modulating Notch activity.
|Rusu, A. D.,
Cornhill, Z. E., Coutiño, B. C., Uribe, M. C., Lourdusamy, A., Markus,
Z., May, S. T., Rahman, R. and Georgiou, M. (2021). CG7379 and ING1 suppress cancer cell invasion by maintaining cell-cell junction integrity. Open Biol 11(9): 210077. PubMed ID: 34493070
Approximately 90% of cancer-related deaths can be attributed to a tumour's ability to spread. This study has identified CG7379, the fly orthologue of human ING1, as a potent invasion suppressor. ING1 is a type II tumour suppressor with well-established roles in the transcriptional regulation of genes that control cell proliferation, response to DNA damage, oncogene-induced senescence and apoptosis. Recent work suggests a possible role for ING1 in cancer cell invasion and metastasis, but the molecular mechanism underlying this observation is lacking. The current results show that reduced expression of CG7379 promotes invasion in vivo in Drosophila, reduces the junctional localization of several adherens and septate junction components, and severely disrupts cell-cell junction architecture. Similarly, ING1 knockdown significantly enhances invasion in vitro and disrupts E-cadherin distribution at cell-cell junctions. A transcriptome analysis reveals that loss of ING1 affects the expression of several junctional and cytoskeletal modulators, confirming ING1 as an invasion suppressor and a key regulator of cell-cell junction integrity.
Thursday, September 23rd - Disease Models
|Olsen, A. L. and Feany, M. B. (2021). Parkinson's disease risk genes act in glia to control neuronal alpha-synuclein toxicity. Neurobiol Dis 159: 105482. PubMed ID: 34390834
Idiopathic Parkinson's disease is the second most common neurodegenerative disease and is estimated to be approximately 30% heritable. Genome wide association studies have revealed numerous loci associated with risk of development of Parkinson's disease. The majority of genes identified in these studies are expressed in glia at either similar or greater levels than their expression in neurons, suggesting that glia may play a role in Parkinson's disease pathogenesis. The role of individual glial risk genes in Parkinson's disease development or progression is unknown, however. It was hypothesized that some Parkinson's disease risk genes exert their effects through glia. A Drosophila model of α-synucleinopathy was developed in which gene expression can be individually expressed in neurons and glia. Human wild type α-synuclein is expressed in all neurons, and these flies develop the hallmarks of Parkinson's disease, including motor impairment, death of dopaminergic and other neurons, and α-synuclein aggregation. In these flies, a candidate genetic screen was performed, using RNAi to knockdown 14 well-validated Parkinson's disease risk genes in glia, and the effect on locomotion was measured in order to identify glial modifiers of the &alpha-synuclein phenotype. Four modifiers were identified: aux, Lrrk, Ric, and Vps13, orthologs of the human genes GAK, LRRK2, RIT2, and VPS13C, respectively. Knockdown of each gene exacerbated neurodegeneration as measured by total and dopaminergic neuron loss. Knockdown of each modifier also increased α-synuclein oligomerization. These results suggest that some Parkinson's disease risk genes exert their effects in glia and that glia can influence neuronal α-synuclein proteostasis in a non-cell-autonomous fashion. Further, this study provides proof of concept that this novel Drosophila α-synucleinopathy model can be used to study glial modifier genes, paving the way for future large unbiased screens to identify novel glial risk factors that contribute to PD risk and progression.
|Krzystek, T. J., Banerjee, R., Thurston, L., Huang, J., Swinter, K., Rahman, S. N., Falzone, T. L. and Gunawardena, S. (2021). Differential mitochondrial roles for alpha-synuclein in DRP1-dependent fission and PINK1/Parkin-mediated oxidation. Cell Death Dis 12(9): 796. PubMed ID: 34404758
Mitochondria are highly dynamic organelles with strict quality control processes that maintain cellular homeostasis. Within axons, coordinated cycles of fission-fusion mediated by dynamin related GTPase protein (DRP1) and mitofusins (MFN), together with regulated motility of healthy mitochondria anterogradely and damaged/oxidized mitochondria retrogradely, control mitochondrial shape, distribution and size. This study has isolated the mechanistic role of α-syn in mitochondrial homeostasis in vivo in a humanized Drosophila model of Parkinson's disease (PD). It was shown that excess α-syn causes fragmented mitochondria, which persists with either truncation of the C-terminus (α(1-120)) or deletion of the NAC region (α(ΔNAC)). Using in vivo oxidation reporters Mito-roGFP2-ORP1/GRX1 and MitoTimer, it was found that α-mediated fragments were oxidized/damaged, but α(1-120)-induced fragments were healthy, suggesting that the C-terminus is required for oxidation. α-mediated oxidized fragments showed biased retrograde motility, but α(1-120)-mediated healthy fragments did not, demonstrating that the C-terminus likely mediates the retrograde motility of oxidized mitochondria. Depletion/inhibition or excess DRP1-rescued α-syn-mediated fragmentation, oxidation, and the biased retrograde motility, indicating that DRP1-mediated fragmentation is likely upstream of oxidation and motility changes. Further, excess PINK/Parkin, two PD-associated proteins that function to coordinate mitochondrial turnover via induction of selective mitophagy, rescued α-syn-mediated membrane depolarization, oxidation and cell death in a C-terminus-dependent manner, suggesting a functional interaction between &alpha-syn and PINK/Parkin. Taken together, these findings identify distinct roles for α-syn in mitochondrial homeostasis, highlighting a previously unknown pathogenic pathway for the initiation of PD.
|Beaver, M., Karisetty, B. C., Zhang, H., Bhatnagar, A., Armour, E., Parmar, V., Brown, R., Xiang, M. and Elefant, F. (2021). Chromatin
and transcriptomic profiling uncover dysregulation of the Tip60
HAT/HDAC2 epigenomic landscape in the neurodegenerative brain. Epigenetics: 1-22. PubMed ID: 34369292
Disruption of histone acetylation-mediated gene control is a critical step in Alzheimer's Disease (AD), yet chromatin analysis of antagonistic histone acetyltransferases (HATs) and histone deacetylases (HDACs) causing these alterations remains uncharacterized. This study reports the first Tip60 HAT versus HDAC2 chromatin (ChIP-seq) and transcriptional (RNA-seq) profiling study in Drosophila melanogaster brains that model early human AD. Tip60 and HDAC2 were found to be predominantly recruited to identical neuronal genes. Moreover, AD brains exhibit robust genome-wide early alterations that include enhanced HDAC2 and reduced Tip60 binding and transcriptional dysregulation. Orthologous human genes to co-Tip60/HDAC2 D. melanogaster neural targets exhibit conserved disruption patterns in AD patient hippocampi. Notably, this study discovered distinct transcription factor binding sites close or within Tip60/HDAC2 co-peaks in neuronal genes, implicating them in coenzyme recruitment. Increased Tip60 protects against transcriptional dysregulation and enhanced HDAC2 enrichment genome-wide. Tip60 HAT/HDAC2 mediated epigenetic neuronal gene disruption is advocated as a genome-wide initial causal event in AD.
|Loganathan, S., Ball, H. E., Manzo, E. and Zarnescu, D. C. (2021). Measuring Glucose Uptake in Drosophila Models of TDP-43 Proteinopathy. J Vis Exp(174). PubMed ID: 34424253
Als is a neurodegenerative disorder causing progressive muscle weakness and death within 2-5 years following diagnosis. Clinical manifestations include weight loss, dyslipidemia, and hypermetabolism; however, it remains unclear how these relate to motor neuron degeneration. Using a Drosophila model of TDP-43 proteinopathy broad ranging metabolic deficits have been identified. Among these, glycolysis was found to be upregulated and genetic interaction experiments provided evidence for a compensatory neuroprotective mechanism. Indeed, despite upregulation of phosphofructokinase, the rate limiting enzyme in glycolysis, an increase in glycolysis using dietary and genetic manipulations was shown to mitigate locomotor dysfunction and increased lifespan in fly models of TDP-43 proteinopathy. To further investigate the effect on TDP-43 proteinopathy on glycolytic flux in motor neurons, a previously reported genetically encoded, FRET-based sensor, FLII12Pglu-700μδ6, was used. This sensor is comprised of a bacterial glucose-sensing domain and cyan and yellow fluorescent proteins as the FRET pair. Upon glucose binding, the sensor undergoes a conformational change allowing FRET to occur. Using FLII12Pglu-700μδ6, glucose uptake was found to be significantly increased in motor neurons expressing TDP-43(G298S), an ALS causing variant. This study shows how to measure glucose uptake, ex vivo, in larval ventral nerve cord preparations expressing the glucose sensor FLII12Pglu-700µδ6 in the context of TDP-43 proteinopathy. This approach can be used to measure glucose uptake and assess glycolytic flux in different cell types or in the context of various mutations causing ALS and related neurodegenerative disorders.
|Liu, Y., Shen, L., Zhang, Y., Zhao, R., Liu, C., Luo, S., Chen, J., Xia, L., Li, T., Peng, Y. and Xia, K. (2021). Rare NRXN1 missense variants identified in autism interfered protein degradation and Drosophila sleeping. J Psychiatr Res 143: 113-122. PubMed ID: 34487988
NRXN1 is involved in synaptogenesis and has been implicated in Autism spectrum disorders (ASDs). However, many rare inherited missense variants of NRXN1 have not been thoroughly evaluated. In this study, functional analyses in vitro and in Drosophila of three NRXN1 missense mutations, Y282H, L893V, and I1135V identified in ASD patients in a previous study were performed. The results showed these three mutations interfered protein degradation compared with NRXN1-WT protein. Expressing human NRXN1 in Drosophila could lead to abnormal circadian rhythm and sleep behavior, and three mutated proteins caused milder phenotypes, indicating the mutations may change the function of NRXN1 slightly. These findings highlight the functional role of rare NRXN1 missense variants identified in autism patients, and provide clues for a better understanding of the pathogenesis of abnormal circadian rhythm and sleep behavior of other organisms, including humans.
Zavortink, M., Golenkina, S., Froldi, F., Dark, C., Cheung, S., Parker,
B. L., Blazev, R., Bakopoulos, D., Christie, E. L., Wimmer, V. C.,
Duckworth, B. C., Richardson, H. E. and Cheng, L. Y. (2021). Tumor-derived MMPs regulate cachexia in a Drosophila cancer model. Dev Cell. PubMed ID: 34473940
Cachexia, the wasting syndrome commonly observed in advanced cancer patients, accounts for up to one-third of cancer-related mortalities. A Drosophila larval model of organ wasting was established whereby epithelial overgrowth in eye-antennal discs leads to wasting of the adipose tissue and muscles. The wasting is associated with fat-body remodeling and muscle detachment and is dependent on tumor-secreted matrix metalloproteinase 1 (Mmp1). Mmp1 can both modulate TGFβ signaling in the fat body and disrupt basement membrane (BM)/extracellular matrix (ECM) protein localization in both the fat body and the muscle. Inhibition of TGFβ signaling or Mmps in the fat body/muscle using a QF2-QUAS binary expression system rescues muscle wasting in the presence of tumor. Altogether,this study proposes that tumor-derived Mmps are central mediators of organ wasting in cancer cachexia.
Wednesday, September 22nd - Enhancers and gene regulation
|Jia, R., Song, Z., Lin, J., Li, Z., Shan, G. and Huang, C. (2021). Gawky modulates MTF-1-mediated transcription activation and metal discrimination. Nucleic Acids Res 49(11): 6296-6314. PubMed ID: 34107019
Metal-induced genes are usually transcribed at relatively low levels under normal conditions and are rapidly activated by heavy metal stress. Many of these genes respond preferentially to specific metal-stressed conditions. However, the mechanism by which the general transcription machinery discriminates metal stress from normal conditions and the regulation of MTF-1-meditated metal discrimination are poorly characterized. Using a focused RNAi screening in Drosophila Schneider 2 (S2) cells, this study identified a novel activator, the Drosophila gawky, of metal-responsive genes. Depletion of gawky has almost no effect on the basal transcription of the metallothionein (MT) genes, but impairs the metal-induced transcription by inducing the dissociation of MTF-1 from the MT promoters and the deficient nuclear import of MTF-1 under metal-stressed conditions. This suggests that gawky serves as a 'checkpoint' for metal stress and metal-induced transcription. In fact, regular mRNAs are converted into gawky-controlled transcripts if expressed under the control of a metal-responsive promoter, suggesting that whether transcription undergoes gawky-mediated regulation is encrypted therein. Additionally, lack of gawky eliminates the DNA binding bias of MTF-1 and the transcription preference of metal-specific genes. This suggests a combinatorial control of metal discrimination by gawky, MTF-1, and MTF-1 binding sites.
|Poupault, C., Choi, D., Lam-Kamath, K., Dewett, D., Razzaq, A., Bunker, J., Perry, A., Cho, I. and Rister, J. (2021). A combinatorial cis-regulatory logic restricts color-sensing Rhodopsins to specific photoreceptor subsets in Drosophila. PLoS Genet 17(6): e1009613. PubMed ID: 34161320
Color vision in Drosophila melanogaster is based on the expression of five different color-sensing Rhodopsin proteins in distinct subtypes of photoreceptor neurons. Promoter regions of less than 300 base pairs are sufficient to reproduce the unique, photoreceptor subtype-specific rhodopsin expression patterns. It has been proposed that the rhodopsin promoters have a bipartite structure: the distal promoter region directs the highly restricted expression in a specific photoreceptor subtype, while the proximal core promoter region provides general activation in all photoreceptors. To distinguish between these two models, the expression patterns were analyzed of a set of hybrid promoters that combine the distal promoter region of one rhodopsin with the proximal core promoter region of another rhodopsin. It was found that the function of the proximal core promoter regions extends beyond providing general activation: these regions play a previously underappreciated role in generating the non-overlapping expression patterns of the different rhodopsins. Therefore, cis-regulatory motifs in both the distal and the proximal core promoter regions recruit transcription factors that generate the unique rhodopsin patterns in a combinatorial manner. This combinatorial regulatory logic is compared to the regulatory logic of olfactory receptor genes and potential implications for the evolution of rhodopsins is discussed.
|Murgas, L., Contreras-Riquelme, S., Martinez-Hernandez, J. E., Villaman, C., Santibanez, R. and Martin, A. J. M. (2021). Automated generation of context-specific gene regulatory networks with a weighted approach in Drosophila melanogaster. Interface Focus 11(4): 20200076. PubMed ID: 34123358
The regulation of gene expression is a key factor in the development and maintenance of life in all organisms. Even so, little is known at whole genome scale for most genes and contexts. This paper proposes a method, Tool for Weighted Epigenomic Networks in Drosophila melanogaster (Fly T-WEoN), to generate context-specific gene regulatory networks starting from a reference network that contains all known gene regulations in the fly. Unlikely regulations are removed by applying a series of knowledge-based filters. Each of these filters is implemented as an independent module that considers a type of experimental evidence, including DNA methylation, chromatin accessibility, histone modifications and gene expression. Fly T-WEoN is based on heuristic rules that reflect current knowledge on gene regulation in D. melanogaster obtained from the literature. Experimental data files can be generated with several standard procedures and used solely when and if available. Fly T-WEoN is available as a Cytoscape application that permits integration with other tools and facilitates downstream network analysis. This work demonstrate the reliability of the method to then provides a relevant application case of this tool: early development of D. melanogaster.
|Prazak, L., Iwasaki, Y., Kim, A. R., Kozlov, K., King, K. and Gergen, J. P. (2021). A dual role for DNA-binding by Runt in activation and repression of sloppy paired transcription. Mol Biol Cell: mbcE20080509. PubMed ID: 34432496
This work investigates the role of DNA-binding by Runt in regulating the sloppy-paired-1 (slp1) gene, and in particular two distinct cis-regulatory elements that mediate regulation by Runt and other pair-rule transcription factors during Drosophila segmentation. A DNA-binding defective form of Runt was found to br ineffective at repressing both the distal (DESE) and proximal (PESE) early stripe elements of slp1 and is also compromised for DESE-dependent activation. The function of Runt-binding sites in DESE is further investigated using site-specific transgenesis and quantitative imaging techniques. When DESE is tested as an autonomous enhancer, mutagenesis of the Runt sites results in a clear loss of Runt-dependent repression but has little to no effect on Runt-dependent activation. Notably, mutagenesis of these same sites in the context of a reporter gene construct that also contains the PESE enhancer results in a significant reduction of DESE-dependent activation as well as the loss of repression observed for the autonomous mutant DESE enhancer. These results provide strong evidence that DNA-binding by Runt directly contributes to the regulatory interplay of interactions between these two enhancers in the early embryo.
|Patel, A. L., Zhang, L., Keenan, S. E., Rushlow, C. A., Fradin, C. and Shvartsman, S. Y. (2021). Capicua is a fast-acting transcriptional brake. Curr Biol. PubMed ID: 34166605
Even though transcriptional repressors are studied with ever-increasing molecular resolution, the temporal aspects of gene repression remain poorly understood. This study addresses the dynamics of transcriptional repression by Capicua (Cic), which is essential for normal development and is commonly mutated in human cancers and neurodegenerative diseases. The speed limit for Cic-dependent gene repression is reported based on live imaging and optogenetic perturbations in the early Drosophila embryo, where Cic was originally discovered. Measurements of Cic concentration and intranuclear mobility, along with real-time monitoring of the activity of Cic target genes, reveal remarkably fast transcriptional repression within minutes of removing an optogenetic de-repressive signal. In parallel, quantitative analyses of transcriptional bursting of Cic target genes support a repression mechanism providing a fast-acting brake on burst generation. This work sets quantitative constraints on potential mechanisms for gene regulation by Cic.
|Postika, N., Schedl, P., Georgiev, P. and Kyrchanova, O. (2021). Redundant enhancers in the iab-5 domain cooperatively activate Abd-B in the A5 and A6 abdominal segments of Drosophila. Development. PubMed ID: 34473267
The Abdominal-B (Abd-B) gene belongs to Bithorax complex and its expression is controlled by four regulatory domains, iab-5, iab-6, iab-7 and iab-8, each of which is thought to be responsible for directing the expression of Abd-B in one of the abdominal segments from A5 to A8. A variety of experiments have supported the idea that BX-C regulatory domains are functionally autonomous and that each domain is both necessary and sufficient to orchestrate the development of the segment they specify. Unexpectedly, this study discovered that this model does not always hold. Instead, it was found that tissue-specific enhancers located in the iab-5 domain are required for the proper activation of Abd-B not only in A5 but also in A6. These findings indicate that the functioning of the iab-5 and iab-6 domains in development of the adult cuticle A5 and A6 in males fit better with an additive model much like that first envisioned by Ed Lewis.
Tuesday, September 21st - Adult neural development and function
|Ismael, S., Sindi, G., Colvin, R. A. and Lee, D. (2021). Activity-dependent release of phosphorylated human tau from Drosophila neurons in primary culture. J Biol Chem: 101108. PubMed ID: 34473990
Neuronal activity can enhance tau release and, thus accelerate tauopathies. This activity-dependent tau release can be used to study the progression of tau (see Drosophila Tau) pathology in Alzheimer's disease (AD), as hyper-phosphorylated tau is implicated in AD pathogenesis and related tauopathies. However, understanding of the mechanisms that regulate activity-dependent tau release from neurons and the role that tau phosphorylation plays in modulating activity-dependent tau release is still rudimentary. In this study, Drosophila neurons in primary culture expressing human tau (hTau) were used to study activity-dependent tau release. hTau release was markedly increased by 50 mM KCl treatment for 1 hour. A similar level of release was observed using optogenetic techniques where genetically targeted neurons were stimulated for 30 min using blue light (470 nm). These results showed that activity-dependent release of phospho-resistant hTau(S11A) was reduced when compared with wild type hTau. In contrast, release of phospho-mimetic hTau(E14) was increased upon activation. Released hTau was phosphorylated in its proline-rich and C-terminal domains using phosphorylation site-specific tau antibodies (e.g., AT8). Fold changes in detectable levels of total or phosphorylated hTau in cell lysates or following immunopurification from conditioned media (IP-CM) were consistent with preferential release of phosphorylated hTau after light stimulation. This study establishes an excellent model to investigate the mechanism of activity-dependent hTau release and to better understand the role of phosphorylated tau release in the pathogenesis of AD since it relates to alterations in the early stage of neurodegeneration associated with increased neuronal activity.
|Messina, G., Prozzillo, Y., Delle Monache, F., Santopietro, M. V., Atterrato, M. T. and Dimitri, P. (2021). The ATPase SRCAP is associated with the mitotic apparatus, uncovering novel molecular aspects of Floating-Harbor syndrome. BMC Biol 19(1): 184. PubMed ID: 34474679
Floating-Harbor syndrome is a rare genetic disease affecting human development caused by dominant truncating mutations in the SRCAP gene, which encodes the ATPase SRCAP, the core catalytic subunit of the homonymous chromatin-remodeling complex. The main function of the SRCAP complex is to promote the exchange of histone H2A with the H2A.Z variant. According to the canonical role played by the SRCAP protein in epigenetic regulation, the Floating-Harbor syndrome is thought to be a consequence of chromatin perturbations. However, additional potential physiological functions of SRCAP have not been sufficiently explored. This study combined cell biology, reverse genetics, and biochemical approaches to study the subcellular localization of the SRCAP protein and assess its involvement in cell cycle progression in HeLa cells. Surprisingly, SRCAP was found to associate with components of the mitotic apparatus (centrosomes, spindle, midbody), interacts with a plethora of cytokinesis regulators, and positively regulates their recruitment to the midbody. Remarkably, SRCAP depletion perturbs both mitosis and cytokinesis. Similarly, DOM-A, the functional SRCAP orthologue in Drosophila melanogaster, is found at centrosomes and the midbody in Drosophila cells, and its depletion similarly affects both mitosis and cytokinesis. These findings provide first evidence suggesting that SRCAP plays previously undetected and evolutionarily conserved roles in cell division, independent of its functions in chromatin regulation. SRCAP may participate in two different steps of cell division: by ensuring proper chromosome segregation during mitosis and midbody function during cytokinesis. Moreover, these findings emphasize a surprising scenario whereby alterations in cell division produced by SRCAP mutations may contribute to the onset of Floating-Harbor syndrome.
|Karekar, P., Jensen,
H. N., Russart, K. L. G., Ponnalagu, D., Seeley, S., Sanghvi, S.,
Smith, S. A., Pyter, L. M., Singh, H. and Gururaja Rao, S. (2021). Tumor-Induced Cardiac Dysfunction: A Potential Role of ROS. Antioxidants (Basel) 10(8). PubMed ID: 34439547
Cancer and heart diseases are the two leading causes of mortality and morbidity worldwide. Many cancer patients undergo heart-related complications resulting in high incidences of mortality. It is generally hypothesized that cardiac dysfunction in cancer patients occurs due to cardiotoxicity induced by therapeutic agents, used to treat cancers and/or cancer-induced cachexia. However, it is not known if localized tumors or unregulated cell growth systemically affect heart function before treatment, and/or prior to the onset of cachexia, hence, making the heart vulnerable to structural or functional abnormalities in later stages of the disease. This study incorporated complementary mouse and Drosophila models to establish if tumor induction indeed causes cardiac defects even before intervention with chemotherapy or onset of cachexia. Focus was placed on one of the key pathways involved in irregular cell growth, the Hippo-Yorkie (Yki) pathway. The transcriptional co-activator of the Yki signaling pathway was overexpressed to induce cellular overgrowth; Yki overexpression in the eye tissue of Drosophila results in compromised cardiac function. These cardiac phenotypes were rescued using antioxidant treatment, with which it is concluded that the Yki induced tumorigenesis causes a systemic increase in ROS affecting cardiac function. These results show that systemic cardiac dysfunction occurs due to abnormal cellular overgrowth or cancer elsewhere in the body; identification of specific cardiac defects associated with oncogenic pathways can facilitate the possible early diagnosis of cardiac dysfunction.
|Guo, Q., Wang, B., Wang, X., Smith, W. W., Zhu, Y. and Liu, Z. (2021). Activation
of Nrf2 in Astrocytes Suppressed PD-Like Phenotypes via Antioxidant and
Autophagy Pathways in Rat and Drosophila Models. Cells 10(8). PubMed ID: 34440619
The oxidative-stress-induced impairment of autophagy plays a critical role in the pathogenesis of Parkinson's disease (PD). This study investigated whether the alteration of Nrf2 in astrocytes protected against 6-OHDA (6-hydroxydopamine)- and rotenone-induced PD-like phenotypes, using 6-OHDA-induced rat PD and rotenone-induced Drosophila PD models. In the PD rat model, Nrf2 expression was significantly higher in astrocytes than in neurons. CDDO-Me (CDDO methyl ester, an Nrf2 inducer) administration attenuated PD-like neurodegeneration mainly through Nrf2 activation in astrocytes by activating the antioxidant signaling pathway and enhancing autophagy in the substantia nigra and striatum. In the PD Drosophila model, the overexpression of Nrf2 in glial cells displayed more protective effects than such overexpression in neurons. Increased Nrf2 expression in glial cells significantly reduced oxidative stress and enhanced autophagy in the brain tissue. The administration of the Nrf2 inhibitor ML385 reduced the neuroprotective effect of Nrf2 through the inhibition of the antioxidant signaling pathway and autophagy pathway. The autophagy inhibitor 3-MA partially reduced the neuroprotective effect of Nrf2 through the inhibition of the autophagy pathway, but not the antioxidant signaling pathway. Moreover, Nrf2 knockdown caused neurodegeneration in flies. Treatment with CDDO-Me attenuated the Nrf2-knockdown-induced degeneration in the flies through the activation of the antioxidant signaling pathway and increased autophagy. An autophagy inducer, rapamycin, partially rescued the neurodegeneration in Nrf2-knockdown Drosophila by enhancing autophagy. These results indicate that the activation of the Nrf2-linked signaling pathways in glial cells plays an important neuroprotective role in PD models.
|Hunt, L. C. and Demontis, F. (2021). Age-Related Increase in Lactate Dehydrogenase Activity in Skeletal Muscle Reduces Lifespan in Drosophila. J Gerontol A Biol Sci Med Sci. PubMed ID: 34477202
Metabolic adaptations occur with aging but the significance and causal roles of such changes are only partially known. In Drosophila, this study found that skeletal muscle aging is paradoxically characterized by increased readouts of glycolysis (lactate, NADH/NAD+) but reduced expression of most glycolytic enzymes. This conundrum is explained by lactate dehydrogenase (LDH), an enzyme necessary for anaerobic glycolysis and whose expression increases with aging. Experimental Ldh overexpression in skeletal muscle of young flies increases glycolysis and shortens lifespan, suggesting that age-related increases in muscle LDH contribute to mortality. Similar results are also found with overexpression of other glycolytic enzymes (Pfrx/PFKFB, Pgi/GPI). Conversely, hypomorphic mutations in Ldh extend lifespan whereas reduction in PFK, Pglym78/PGAM, Pgi/GPI, and Ald/ALDO levels shorten lifespan to various degrees, indicating that glycolysis needs to be tightly controlled for optimal aging. Altogether, these findings indicate a role for muscle LDH and glycolysis in aging.
|Jang, H. J.,
Le, M. U. T., Park, J. H., Chung, C. G., Shon, J. G., Lee, G. S., Moon,
J. H., Lee, S. B., Choi, J. S., Lee, T. G. and Yoon, S. (2021). Matrix-Assisted
Laser Desorption/Ionization Mass Spectrometry Imaging of Phospholipid
Changes in a Drosophila Model of Early Amyotrophic Lateral Sclerosis. J Am Soc Mass Spectrom. PubMed ID: 34448582
ALS is a degenerative disease caused by motor neuron damage in the central nervous system. Drosophila is widely used to investigate disease mechanisms. MALDI-MSI was performed to investigate changes in phospholipid distribution in the brain tissue of an ALS-induced Drosophila model. Fly brain tissues of several hundred micrometers or less were sampled using a fly collar to obtain reproducible tissue sections of similar sizes. MSI of brain tissues of Drosophila cultured for 1 or 10 days showed that the distribution of phospholipids, including phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidic acid (PA), phosphatidylserine (PS), and phosphatidylinositol (PI), was significantly different between the control group and the ALS group. In addition, the lipid profile according to phospholipids differed as the culture time increased from 1 to 10 days. These results suggest that disease indicators based on lipid metabolites can be discovered by performing MALDI-MSI on very small brain tissue samples from the Drosophila disease model to ultimately assess the phospholipid changes that occur in early-stage ALS.
Friday September 17th - Immune Response
|Hanson, M. A., Cohen, L. B., Marra, A., Iatsenko, I., Wasserman, S. A. and Lemaitre, B. (2021). The Drosophila Baramicin polypeptide gene protects against fungal infection. PLoS Pathog 17(8): e1009846. PubMed ID: 34432851
The fruit fly Drosophila melanogaster combats microbial infection by producing a battery of effector peptides that are secreted into the haemolymph. Technical difficulties prevented the investigation of these short effector genes until the recent advent of the CRISPR/CAS era. As a consequence, many putative immune effectors remain to be formally described, and exactly how each of these effectors contribute to survival is not well characterized. This study describes a novel Drosophila antifungal peptide gene that was named Baramicin A. BaraA was shown to encodes a precursor protein cleaved into multiple peptides via furin cleavage sites. BaraA is strongly immune-induced in the fat body downstream of the Toll pathway, but also exhibits expression in other tissues. Importantly, it was shown that flies lacking BaraA are viable but susceptible to the entomopathogenic fungus Beauveria bassiana. Consistent with BaraA being directly antimicrobial, overexpression of BaraA promotes resistance to fungi and the IM10-like peptides produced by BaraA synergistically inhibit growth of fungi in vitro when combined with a membrane-disrupting antifungal. Surprisingly, BaraA mutant males but not females display an erect wing phenotype upon infection. This study has characterized a new antifungal immune effector downstream of Toll signalling, and show it is a key contributor to the Drosophila antimicrobial response.
|Bhattacharya, T., Rice, D. W., Crawford, J. M., Hardy, R. W. and Newton, I. L. G. (2021). Evidence
of Adaptive Evolution in Wolbachia-Regulated Gene DNMT2 and Its Role in
the Dipteran Immune Response and Pathogen Blocking. Viruses 13(8). PubMed ID: 34452330
Eukaryotic nucleic acid methyltransferase (MTase) proteins are essential mediators of epigenetic and epitranscriptomic regulation. DNMT2 belongs to a large, conserved family of DNA MTases found in many organisms, including holometabolous insects such as fruit flies and mosquitoes, where it is the lone MTase. Interestingly, despite its nomenclature, DNMT2 is not a DNA MTase, but instead targets and methylates RNA species. A growing body of literature suggests that DNMT2 mediates the host immune response against a wide range of pathogens, including RNA viruses. Curiously, although DNMT2 is antiviral in Drosophila, its expression promotes virus replication in mosquito species. Therefore this study sought to understand the divergent regulation, function, and evolution of these orthologs. The role is described of the Drosophila-specific host protein IPOD in regulating the expression and function of fruit fly DNMT2. Heterologous expression of these orthologs suggests that DNMT2's role as an antiviral is host-dependent, indicating a requirement for additional host-specific factors. Finally, this study identified and describes potential evidence of positive selection at different times throughout DNMT2 evolution within dipteran insects. Specific codons within each ortholog were identified that are under positive selection, and they were found to be restricted to four distinct protein domains, which likely influence substrate binding, target recognition, and adaptation of unique intermolecular interactions. Collectively, these findings highlight the evolution of DNMT2 in Dipteran insects and point to structural, regulatory, and functional differences between mosquito and fruit fly homologs.
Winther, K. G., Gad, H. H., Ai, X., Chen, Y., Li, L., Wei, Z., Deng, H.,
Liu, J., Frederiksen, N. A., Simonsen, B., Andersen, L. L., Kleigrewe,
K., Dalskov, L., Pichlmair, A., Cai, H., Imler, J. L. and Hartmann, R.
(2021). Two cGAS-like receptors induce antiviral immunity in Drosophila. Nature. PubMed ID: 34261128
In mammals, cyclic GMP-AMP (cGAMP) synthase (cGAS) produces the cyclic dinucleotide (CDN) 2'3'-cGAMP in response to cytosolic DNA and this triggers an antiviral immune response. cGAS belongs to a large family of cGAS/DncV-like nucleotidyltransferases, present in both prokaryotes and eukaryotes. In bacteria, these enzymes synthesize a range of cyclic oligonucleotide and have recently emerged as important regulators of phage infections. This study identified two novel cGAS-like receptors (cGLRs) in the insect Drosophila melanogaster. cGLR1 and cGLR2 activate Sting and NF-κB dependent antiviral immunity in response to infection with RNA or DNA viruses. cGLR1 is activated by dsRNA to produce the novel CDN 3'2'-cGAMP whereas cGLR2 produces a combination of 2'3'-cGAMP and 3'2' cGAMP in response to a yet unidentified stimulus. These data establish cGAS as the founding member of a family of receptors sensing different types of nucleic acids and triggering immunity through production of CDNs beyond 2'3'-cGAMP.
|Salem Wehbe, L., Barakat, D., Acker, A., El Khoury, R., Reichhart, J. M., Matt, N. and El Chamy, L. (2021). Protein Phosphatase 4 Negatively Regulates the Immune Deficiency-NF-kappaB Pathway during the Drosophila Immune Response. J Immunol 207(6): 1616-1626. PubMed ID: 34452932
The evolutionarily conserved immune deficiency (IMD) signaling pathway shields Drosophila against bacterial infections. It regulates the expression of antimicrobial peptides encoding genes through the activation of the NF-κB transcription factor Relish. Tight regulation of the signaling cascade ensures a balanced immune response, which is otherwise highly harmful. Several phosphorylation events mediate intracellular progression of the IMD pathway. However, signal termination by dephosphorylation remains largely elusive. This study identifyied the highly conserved protein phosphatase 4 (PP4) complex as a bona fide negative regulator of the IMD pathway. RNA interference-mediated gene silencing of PP4-19c, PP4R2, and Falafel, which encode the catalytic and regulatory subunits of the phosphatase complex, respectively, caused a marked upregulation of bacterial-induced antimicrobial peptide gene expression in both Drosophila melanogaster S2 cells and adult flies. Deregulated IMD signaling is associated with reduced lifespan of PP4-deficient flies in the absence of any infection. In contrast, flies overexpressing this phosphatase are highly sensitive to bacterial infections. Altogether, these results highlight an evolutionarily conserved function of PP4c in the regulation of NF-κB signaling from Drosophila to mammals.
|Jugder, B. E., Kamareddine, L. and Watnick, P. I. (2021). Microbiota-derived acetate activates intestinal innate immunity via the Tip60 histone acetyltransferase complex. Immunity. PubMed ID: 34107298
Microbe-derived acetate activates the Drosophila immunodeficiency (IMD) pathway in a subset of enteroendocrine cells (EECs) of the anterior midgut. In these cells, the IMD pathway co-regulates expression of antimicrobial and enteroendocrine peptides including tachykinin, a repressor of intestinal lipid synthesis. To determine whether acetate acts on a cell surface pattern recognition receptor or an intracellular target, it was asked whether acetate import was essential for IMD signaling. Mutagenesis and RNA interference revealed that the putative monocarboxylic acid transporter Tarag was essential for enhancement of IMD signaling by dietary acetate. Interference with histone deacetylation in EECs augmented transcription of genes regulated by the steroid hormone ecdysone including IMD targets. Reduced expression of the histone acetyltransferase Tip60 decreased IMD signaling and blocked rescue by dietary acetate and other sources of intracellular acetyl-CoA. Thus, microbe-derived acetate induces chromatin remodeling within enteroendocrine cells, co-regulating host metabolism and intestinal innate immunity via a Tip60-steroid hormone axis that is conserved in mammals.
|Zhou, J., Valentini, E. and Boutros, M. (2021). Microenvironmental innate immune signaling and cell mechanical responses promote tumor growth. Dev Cell 56(13): 1884-1899. PubMed ID: 34197724
Tissue homeostasis is achieved by balancing stem cell maintenance, cell proliferation and differentiation, as well as the purging of damaged cells. Elimination of unfit cells maintains tissue health; however, the underlying mechanisms driving competitive growth when homeostasis fails, for example, during tumorigenesis, remain largely unresolved. Using a Drosophila intestinal model, this study found that tumor cells outcompete nearby enterocytes (ECs) by influencing cell adhesion and contractility. This process relies on activating the immune-responsive Relish/NF-κB pathway to induce EC delamination and requires a JNK-dependent transcriptional upregulation of the peptidoglycan recognition protein PGRP-LA. Consequently, in organisms with impaired PGRP-LA function, tumor growth is delayed and lifespan extended. This study identifies a non-cell-autonomous role for a JNK/PGRP-LA/Relish signaling axis in mediating death of neighboring normal cells to facilitate tumor growth. It is proposed that intestinal tumors "hijack" innate immune signaling to eliminate enterocytes in order to support their own growth.
Thursday September 16th - RNA and Transposons
|Chen, P., Luo, Y. and Aravin, A. A. (2021). RDC complex executes a dynamic piRNA program during Drosophila spermatogenesis to safeguard male fertility. PLoS Genet 17(9): e1009591. PubMed ID: 34473737
piRNAs are small non-coding RNAs that guide the silencing of transposons and other targets in animal gonads. In Drosophila female germline, many piRNA source loci dubbed "piRNA clusters" lack hallmarks of active genes and exploit an alternative path for transcription, which relies on the Rhino-Deadlock-Cutoff (RDC) complex. RDC was thought to be absent in testis, so it remains to date unknown how piRNA cluster transcription is regulated in the male germline. This study found that components of RDC complex are expressed in male germ cells during early spermatogenesis, from germline stem cells (GSCs) to early spermatocytes. RDC is essential for expression of dual-strand piRNA clusters and transposon silencing in testis; however, it is dispensable for expression of Y-linked Suppressor of Stellate piRNAs and therefore Stellate silencing. Despite intact Stellate repression, males lacking RDC exhibited compromised fertility accompanied by germline DNA damage and GSC loss. Thus, piRNA-guided repression is essential for normal spermatogenesis beyond Stellate silencing. While RDC associates with multiple piRNA clusters in GSCs and early spermatogonia, its localization changes in later stages as RDC concentrates on a single X-linked locus, AT-chX. Dynamic RDC localization is paralleled by changes in piRNA cluster expression, indicating that RDC executes a fluid piRNA program during different stages of spermatogenesis. These results disprove the common belief that RDC is dispensable for piRNA biogenesis in testis and uncover the unexpected, sexually dimorphic and dynamic behavior of a core piRNA pathway machinery.
|Klann, M., Issa, A. R., Pinho, S. and Alonso, C. R. (2021). microRNA-dependent control of sensory neuron function regulates posture behaviour in Drosophila. J Neurosci. PubMed ID: 34417328
Sensory neuronsrepresent a critical component in all neural circuits and their correct function is essential for the generation of behaviour and adaptation to the environment. This study reports that the evolutionarily-conserved microRNA (miRNA) miR-263b, plays a key behavioural role in Drosophila melanogaster through effects on the function of larval sensory neurons. Several independent experiments (in 50:50/male:female populations) support this finding: first, miRNA expression analysis - via reporter expression and FACS-qPCR analysis - demonstrate miR-263b expression in larval sensory neurons. Second, behavioural tests in miR-263b null mutants show defects in self-righting, an innate and evolutionarily conserved posture-control behaviour that allows larvae to rectify their position if turned upside-down. Third, competitive inhibition of miR-263b in sensory neurons using a miR-263b 'sponge' leads to self-righting defects. Fourth, systematic analysis of sensory neurons in miR-263b mutants shows no detectable morphological defects in their stereotypic pattern, whilst genetically-encoded calcium sensors expressed in the sensory domain reveal a reduction in neural activity in miR-263b mutants. Fifth, miR-263b null mutants show reduced 'touch-response' behaviour and a compromised response to sound, both characteristic of larval sensory deficits. Furthermore, bioinformatic miRNA target analysis, gene expression assays, and behavioural phenocopy experiments suggest that miR-263b might exert its effects - at least in part - through repression of the bHLH transcription factor atonal. Altogether, this study suggests a model in which miRNA-dependent control of transcription factor expression affects sensory function and behaviour.
|Kelleher, E. S. (2021). Protein-Protein Interactions Shape Genomic Autoimmunity in the Adaptively Evolving Rhino-Deadlock-Cutoff Complex. Genome Biol Evol 13(7). PubMed ID: 34115120
The Piwi-interacting RNA (piRNA) pathway is a genomic defense system that controls the movement of transposable elements (TEs) through transcriptional and post-transcriptional silencing. Although TE defense is critical to ensuring germline genome integrity, it is equally critical that the piRNA pathway avoids autoimmunity in the form of silencing host genes. Ongoing cycles of selection for expanded control of invading TEs, followed by selection for increased specificity to reduce impacts on host genes, are proposed to explain the frequent signatures of adaptive evolution among piRNA pathway proteins. However, empirical tests of this model remain limited, particularly with regards to selection against genomic autoimmunity. This study examined three adaptively evolving piRNA proteins, Rhino, Deadlock, and Cutoff, for evidence of interspecific divergence in autoimmunity between Drosophila melanogaster and Drosophila simulans. This study tested key prediction of the autoimmunity hypothesis that foreign heterospecific piRNA proteins will exhibit enhanced autoimmunity, due to the absence of historical selection against off-target effects. Consistent with this prediction, full-length D. simulans Cutoff, as well as the D. simulans hinge and chromo domains of Rhino, exhibit expanded regulation of D. melanogaster genes. It was further demonstrated that this autoimmunity is dependent on known incompatibilities between D. simulans proteins or domains and their interacting partners in D. melanogaster. These observations reveal that the same protein-protein interaction domains that are interfaces of adaptive evolution in Rhino and Cutoff also determine their potential for autoimmunity.
|Zhu, L. and Fukunaga, R. (2021). RNA-binding
protein Maca is crucial for gigantic male fertility factor gene
expression, spermatogenesis, and male fertility, in Drosophila. PLoS Genet 17(6): e1009655. PubMed ID: 34181646
During spermatogenesis, the process in which sperm for fertilization are produced from germline cells, gene expression is spatiotemporally highly regulated. In Drosophila, successful expression of extremely large male fertility factor genes on Y-chromosome spanning some megabases due to their gigantic intron sizes is crucial for spermatogenesis. Expression of such extremely large genes must be challenging, but the molecular mechanism that allows it remains unknown. This study reports that a novel RNA-binding protein Maca, which contains two RNA-recognition motifs, is crucial for this process. maca null mutant male flies exhibited a failure in the spermatid individualization process during spermatogenesis, lacked mature sperm, and were completely sterile, while maca mutant female flies were fully fertile. Proteomics and transcriptome analyses revealed that both protein and mRNA abundance of the gigantic male fertility factor genes kl-2, kl-3, and kl-5 (kl genes) are significantly decreased, where the decreases of kl-2 are particularly dramatic, in maca mutant testes. Splicing of the kl-3 transcripts was also dysregulated in maca mutant testes. All these physiological and molecular phenotypes were rescued by a maca transgene in the maca mutant background. Furthermore, it was found that in the control genetic background, Maca is exclusively expressed in spermatocytes in testes and enriched at Y-loop A/C in the nucleus, where the kl-5 primary transcripts are localized. These data suggest that Maca increases transcription processivity, promotes successful splicing of gigantic introns, and/or protects transcripts from premature degradation, of the kl genes. This study identified a novel RNA-binding protein Maca that is crucial for successful expression of the gigantic male fertility factor genes, spermatogenesis, and male fertility.
|Nguyen, D., Buisine, N., Fayol, O., Michels, A. A., Bensaude, O., Price, D. H. and Uguen, P. (2021). An alternative D. melanogaster 7SK snRNPd. BMC Mol Cell Biol 22(1): 43. PubMed ID: 34461828
The 7SK small nuclear RNA (snRNA) found in most metazoans is a key regulator of P-TEFb which in turn regulates RNA polymerase II elongation. Although its primary sequence varies in protostomes, its secondary structure and function are conserved across evolutionary distant taxa. This study describes a novel ncRNA sharing many features characteristic of 7SK RNAs, in D. melanogaster. Yhe structure of the corresponding gene was examined and the expression profiles were determined of the encoded RNA, called snRNA:7SK:94F, during development. It is probably produced from the transcription of a lncRNA which is processed into a mature snRNA. This study also addressed its biological function and showed that, like dm7SK, this alternative 7SK interacts in vivo with the different partners of the P-TEFb complex, i.e. HEXIM, LARP7 and Cyclin T. This novel RNA is widely expressed across tissues. It is proposed that two distinct 7SK genes might contribute to the formation of the 7SK snRNP complex in D. melanogaster.
|Zhang, J., Wen, D., Li, E. Y., Palli, S. R., Li, S., Wang, J. and Liu, S. (2021). MicroRNA
miR-8 promotes cell growth of corpus allatum and juvenile hormone
biosynthesis independent of insulin/IGF signaling in Drosophila
melanogaster. Insect Biochem Mol Biol 136: 103611. PubMed ID: 34182107
The Drosophila melanogaster corpus allatum (CA) produces and releases three types of sesquiterpenoid hormones, including juvenile hormone III bisepoxide (JHB3), juvenile hormone III (JH III), and methyl farnesoate (MF). JH biosynthesis involves multiple discrete enzymatic reactions and is subjected to a comprehensive regulatory network including microRNAs (miRNAs). Using a high throughput sequencing approach, abundant miRNAs were identified in the D. melanogaster ring gland, which consists of the CA, prothoracic gland, and corpus cardiaca. miR-8 was identified as a potential candidate for regulation of metamorphosis and its role in the CA was further studied. Overexpression of miR-8 in the CA increased cell size of the gland and expression of Jhamt (a gene coding for a key regulatory enzyme in JH biosynthesis), resulting in pupal lethality. By contrast, sponge-mediated reduction of miR-8 in the CA decreased cell size and Jhamt expression, but did not cause lethality. Further investigation revealed that miR-8 promotes cell growth independent of insulin/IGF signaling. Taken together, these experiments show that miR-8 is highly expressed in the CA and exerts its positive effects on cell growth and JH biosynthesis.
Wednesday, September 15th - Signaling
|Fagan, R. R., Kearney, P. J., Luethi, D., Bolden, N. C., Sitte, H. H., Emery, P. and Melikian, H. E. (2021). Dopaminergic
Ric GTPase activity impacts amphetamine sensitivity and sleep quality
in a dopamine transporter-dependent manner in Drosophila melanogaster. Mol Psychiatry. PubMed ID: 34471250
Dopamine (DA) is required for movement, sleep, and reward, and DA signaling is tightly controlled by the presynaptic DA transporter (DAT). Therapeutic and addictive psychostimulants, including methylphenidate (Ritalin; MPH), cocaine, and amphetamine (AMPH), markedly elevate extracellular DA via their actions as competitive DAT inhibitors (MPH, cocaine) and substrates (AMPH). DAT silencing in mice and invertebrates results in hyperactivity, reduced sleep, and blunted psychostimulant responses, highlighting DAT's essential role in DA-dependent behaviors. DAT surface expression is not static; rather it is dynamically regulated by endocytic trafficking. PKC-stimulated DAT endocytosis requires the neuronal GTPase, Rit2, and Rit2 silencing in mouse DA neurons impacts psychostimulant sensitivity. However, it is unknown whether or not Rit2-mediated changes in psychostimulant sensitivity are DAT-dependent. This study leveraged Drosophila melanogaster to test whether the Drosophila Rit2 ortholog, Ric, impacts dDAT function, trafficking, and DA-dependent behaviors. Orthologous to hDAT and Rit2, dDAT and Ric directly interact, and the constitutively active Ric mutant Q117L increased dDAT surface levels and function in cell lines and ex vivo Drosophila brains. Moreover, DAergic RicQ117L expression caused sleep fragmentation in a DAT-dependent manner but had no effect on total sleep and daily locomotor activity. Importantly, this study found that Rit2 is required for AMPH-stimulated DAT internalization in mouse striatum, and that DAergic RicQ117L expression significantly increased Drosophila AMPH sensitivity in a DAT-dependent manner, suggesting a conserved impact of Ric-dependent DAT trafficking on AMPH sensitivity. These studies support that the DAT/Rit2 interaction impacts both baseline behaviors and AMPH sensitivity, potentially by regulating DAT trafficking.
|Ganguly, P., Madonsela, L., Chao, J. T., Loewen, C. J. R., O'Connor, T. P., Verheyen, E. M. and Allan, D. W. (2021). A
scalable Drosophila assay for clinical interpretation of human PTEN
variants in suppression of PI3K/AKT induced cellular proliferation. PLoS Genet 17(9): e1009774. PubMed ID: 34492006
Gene variant discovery is becoming routine, but it remains difficult to usefully interpret the functional consequence or disease relevance of most variants. To fill this interpretation gap, experimental assays of variant function are becoming common place. Yet, it remains challenging to make these assays reproducible, scalable to high numbers of variants, and capable of assessing defined gene-disease mechanism for clinical interpretation aligned to the ClinGen Sequence Variant Interpretation (SVI) Working Group guidelines for 'well-established assays'. Drosophila melanogaster offers great potential as an assay platform, but was untested for high numbers of human variants adherent to these guidelines. This study tested the utility of Drosophila as a platform for scalable well-established assays. A genetic interaction approach was taken to test the function of ~100 human PTEN (see Drosophila Pten) variants in cancer-relevant suppression of PI3K/AKT signaling in cellular growth and proliferation. The assay was validated using biochemically characterized PTEN mutants as well as 23 total known pathogenic and benign PTEN variants, all of which the assay correctly assigned into predicted functional categories. Additionally, function calls for these variants correlated very well with recent published data from a human cell line. Finally, using these pathogenic and benign variants to calibrate the assay, readout thresholds could be set for clinical interpretation of the pathogenicity of 70 other PTEN variants. Overall, this study demonstrated that Drosophila offers a powerful assay platform for clinical variant interpretation, that can be used in conjunction with other well-established assays, to increase confidence in the accurate assessment of variant function and pathogenicity.
|Ding, G., Xiang, X.,
Hu, Y., Xiao, G., Chen, Y., Binari, R., Comjean, A., Li, J., Rushworth,
E., Fu, Z., Mohr, S. E., Perrimon, N. and Song, W. (2021). Coordination of tumor growth and host wasting by tumor-derived Upd3. Cell Rep 36(7): 109553. PubMed ID: 34407411
yki-induced gut tumors in Drosophila are associated with host wasting, including muscle dysfunction, lipid loss, and hyperglycemia, a condition reminiscent of human cancer cachexia. This model has been used to identify tumor-derived ligands that contribute to host wasting. To identify additional molecular networks involved in host-tumor interactions, PathON, a web-based tool analyzing the major signaling pathways in Drosophila was developed, and the Upd3/Jak/Stat axis was uncovered as an important modulator. yki-gut tumors were found to secrete Upd3 to promote self-overproliferation and enhance Jak/Stat signaling in host organs to cause wasting, including muscle dysfunction, lipid loss, and hyperglycemia. It was further revealed that Upd3/Jak/Stat signaling in the host organs directly triggers the expression of ImpL2, an antagonistic binding protein for insulin-like peptides, to impair insulin signaling and energy balance. Altogether, these results demonstrate that yki-gut tumors produce a Jak/Stat pathway ligand, Upd3, that regulates both self-growth and host wasting.
|Duan, X., Xu, L., Li, Y., Jia, L., Liu, W., Shao, W., Bayat, V., Shang, W., Wang, L., Liu, J. P. and Tong, C. (2021). Regulation of lipid homeostasis by the TBC protein dTBC1D22 via modulation of the small GTPase Rab40 to facilitate lipophagy. Cell Rep 36(9): 109541. PubMed ID: 34469730
The regulation of lipid homeostasis is not well understood. Using forward genetic screening, this study demonstrates that the loss of dTBC1D22 (CG5745), an essential gene that encodes a Tre2-Bub2-Cdc16 (TBC) domain-containing protein, results in lipid droplet accumulation in multiple tissues. dTBC1D22 interacts with Rab40 and exhibits GTPase activating protein (GAP) activity. Overexpression of either the GTP- or GDP-binding-mimic form of Rab40 results in lipid droplet accumulation. Rab40 mutant flies are defective in lipid mobilization. The lipid depletion induced by overexpression of Brummer, a triglyceride lipase, is dependent on Rab40. Rab40 mutant flies exhibit decreased lipophagy and small size of autolysosomal structures, which may be due to the defective Golgi functions. Finally, it was demonstrated that Rab40 physically interacts with Lamp1, and Rab40 is required for the distribution of Lamp1 during starvation. It is proposed that dTBC1D22 functions as a GAP for Rab40 to regulate lipophagy.
|Ho, K. Y. L., Khadilkar, R. J., Carr, R. L. and Tanentzapf, G. (2021). A gap-junction-mediated, calcium-signaling network controls blood progenitor fate decisions in hematopoiesis. Curr Biol. PubMed ID: 34480855
Stem cell homeostasis requires coordinated fate decisions among stem cells that are often widely distributed within a tissue at varying distances from their stem cell niche. This requires a mechanism to ensure robust fate decisions within a population of stem cells. This study shows that, in the Drosophila hematopoietic organ, the lymph gland (LG), gap junctions (see Drosophila Ogre) form a network that coordinates fate decisions between blood progenitors. Using live imaging of calcium signaling in intact LGs, it was found that blood progenitors are connected through a signaling network. Blocking gap junction function disrupts this network, alters the pattern of encoded calcium signals, and leads to loss of progenitors and precocious blood cell differentiation. Ectopic and uniform activation of the calcium-signaling mediator CaMKII restores progenitor homeostasis when gap junctions are disrupted. Overall, these data show that gap junctions equilibrate cell signals between blood progenitors to coordinate fate decisions and maintain hematopoietic homeostasis.
Ferreira, R. M., Xu, H., Goncalves, M., Barros-Carvalho, A., Cravo, J.,
Maia, A. F., Carneiro, P., Figueiredo, C., Smith, M. L., Stamenovic,
D., Morais-de-Sa, E. and Seruca, R. (2021). Integrin beta1 orchestrates the abnormal cell-matrix attachment and invasive behaviour of E-cadherin dysfunctional cells. Gastric Cancer. PubMed ID: 34486077
Tumour progression relies on the ability of cancer cells to penetrate and invade neighbouring tissues. E-cadherin loss is associated with increased cell invasion in gastric carcinoma. To identify ECM components and receptors relevant for adhesion of E-cadherin dysfunctional cells, a novel ECM microarray platform was implemented coupled with molecular interaction networks. The functional role of putative candidates was determined by combining micropattern traction microscopy, protein modulation and in vivo approaches, as well as transcriptomic data of 262 gastric carcinoma samples, retrieved from the cancer genome atlas (TCGA). This study shows that E-cadherin mutations induce an abnormal interplay of cells with specific components of the ECM, which encompasses increased traction forces and Integrin β1 (see Drosophila Myospheroid) activation. Integrin β1 synergizes with E-cadherin dysfunction, promoting cell scattering and invasion. The significance of the E-cadherin-Integrin β1 crosstalk was validated in Drosophila models. It is concluded that integrin β1 is a key mediator of invasion in carcinomas with E-cadherin impairment and should be regarded as a biomarker of poor prognosis in gastric cancer.
Monday, September 13th - Larval and Adult Behavior
|Guo, D., Zhang, Y.
J., Zhang, S., Li, J., Guo, C., Pan, Y. F., Zhang, N., Liu, C. X., Jia,
Y. L., Li, C. Y., Ma, J. Y., Nassel, D. R., Gao, C. F. and Wu, S. F.
(2021). Cholecystokinin-like peptide mediates satiety by inhibiting sugar attraction. PLoS Genet 17(8): e1009724. PubMed ID: 34398892
Feeding is essential for animal survival and reproduction and is regulated by both internal states and external stimuli. However, little is known about how internal states influence the perception of external sensory cues that regulate feeding behavior. This study investigated the neuronal and molecular mechanisms behind nutritional state-mediated regulation of gustatory perception in control of feeding behavior in the brown planthopper and Drosophila. Feeding was found to increase the expression of the cholecystokinin-like peptide, sulfakinin (SK), and the activity of a set of SK-expressing neurons. Starvation elevates the transcription of the sugar receptor Gr64f and SK negatively regulates the expression of Gr64f in both insects. Interestingly, it was found that one of the two known SK receptors, CCKLR-17D3, is expressed by some of Gr64f-expressing neurons in the proboscis and proleg tarsi. Thus, this study has identified SK as a neuropeptide signal in a neuronal circuitry that responds to food intake, and regulates feeding behavior by diminishing gustatory receptor gene expression and activity of sweet sensing GRNs. The findings demonstrate one nutritional state-dependent pathway that modulates sweet perception and thereby feeding behavior, but the experiments cannot exclude further parallel pathways. Importantly, it was shown that the underlying mechanisms are conserved in the two distantly related insect species.
|Feng, X., Hong, X., Fan, Q., Chen, L., Li, J., Deng, J., Gong, S., Hou, F. F. and Zhang, F. (2021). dCubilin/dAMN- mediated protein reabsorption in Drosophila nephrocytes modulates longevity. Dis Model Mech. PubMed ID: 34437681
Aging is a multi-faceted process regulated by multiple cellular pathways, including the proteostasis network. Pharmacological or genetic enhancement of the intracellular proteostasis network extends lifespan and prevents age-related diseases. However, how proteostasis is regulated in different tissues throughout the aging process remains unclear. This study shows that Drosophila homologs for Cubilin/Amnionless (dCubilin/dAMN)-mediated protein reabsorption from hemolymph (fly equivalent of blood) by nephrocytes modulates longevity through regulating proteostasis in muscle and brain tissues in Drosophila. Overexpression of dAMN receptor in nephrocytes extends lifespan, whereas nephrocyte-specific dCubilin or dAMN RNAi knockdown results in a protein reabsorption defect and shortens lifespan in flies. dCubilin/dAMN-mediated protein reabsorption in nephrocytes regulates proteostasis in hemolymph and improves healthspan. In addition, it was shown that enhanced dCubilin/dAMN-mediated protein reabsorption in nephrocytes slows down the aging process in muscle and brain by maintaining the proteostasis network in these tissues. Furthermore, this study shows that dCubilin/dAMN -mediated protein reabsorption in nephrocytes affects proteasome activity in the whole body and muscle tissues. Altogether, this work has revealed an inter-organ communication network across nephrocytes and muscle/neuronal tissue which is essential to maintain proteostasis and to delay senescence in these organs. The findings have provided insights into the role of renal protein reabsorption in the aging process via this tele-proteostasis network.
Chen, A., Budelli, G., Berck, M. E., Richter, V., Rist, A., Thum, A.
S., Cardona, A., Klein, M., Garrity, P. and Samuel, A. D. T. (2021). Synchronous and opponent thermosensors use flexible cross-inhibition to orchestrate thermal homeostasis. Sci Adv 7(35). PubMed ID: 34452914
Body temperature homeostasis is essential and reliant upon the integration of outputs from multiple classes of cooling- and warming-responsive cells. The computations that integrate these outputs are not understood. This study discovered a set of warming cells (WCs) and show that the outputs of these WCs combine with previously described cooling cells (CCs) in a cross-inhibition computation to drive thermal homeostasis in larval Drosophila WCs and CCs detect temperature changes using overlapping combinations of ionotropic receptors: Ir68a, Ir93a, and Ir25a for WCs and Ir21a, Ir93a, and Ir25a for CCs. WCs mediate avoidance to warming while cross-inhibiting avoidance to cooling, and CCs mediate avoidance to cooling while cross-inhibiting avoidance to warming. Ambient temperature-dependent regulation of the strength of WC- and CC-mediated cross-inhibition keeps larvae near their homeostatic set point. Using neurophysiology, quantitative behavioral analysis, and connectomics, this study demonstrated how flexible integration between warming and cooling pathways can orchestrate homeostatic thermoregulation.
Fanelli, G., Silvestri, F., Cherubini, A., Del Quondam, S., Bongiorni,
S., Taddei, A. R., Ceci, M., De Palma, C., Perrotta, C., Rinalducci, S.,
Prantera, G. and Cervia, D. (2021). Nutraceutical Strategy to
Counteract Eye Neurodegeneration and Oxidative Stress in Drosophila
melanogaster Fed with High-Sugar Diet. Antioxidants (Basel) 10(8). PubMed ID: 34439445
Aberrant production of reactive oxygen species (ROS) is a common feature of damaged retinal neurons in diabetic retinopathy, and antioxidants may exert both preventive and therapeutic action. To evaluate the beneficial and antioxidant properties of food supplementation with Lisosan G, a powder of bran and germ of grain (Triticum aestivum) obtained by fermentation with selected lactobacillus and natural yeast strains, an in vivo model was used of hyperglycemia-induced retinal damage, the fruit fly Drosophila melanogaster fed with high-sucrose diet. Lisosan G positively affected the visual system of hyperglycemic flies at structural/functional level, decreased apoptosis, and reactivated protective autophagy at the retina internal network. Also, in high sucrose-fed Drosophila, Lisosan G reduced the levels of brain ROS and retina peroxynitrite. The analysis of oxidative stress-related metabolites suggested key mediators of Lisosan G-induced inhibition of neuronal ROS, along with the upregulation of glutathione system. Of note, Lisosan G may impact oxidative stress and the ensuing retinal cell death, also independently from autophagy, although the autophagy-ROS cross-talk is critical. This study demonstrates that supplementation with Lisosan G exerts a antioxidant effect on retinal neurons, thus providing efficacious neuroprotection of hyperglycemic eye.
|Akhmetova, K., Balasov, M. and Chesnokov, I. (2021). Drosophila STING protein has a role in lipid metabolism. Elife 10. PubMed ID: 34467853
Stimulator of interferon genes (STING) plays an important role in innate immunity by controlling type I interferon response against invaded pathogens. This work describes a previously unknown role of STING in lipid metabolism in Drosophila. Flies with STING deletion are sensitive to starvation and oxidative stress, have reduced lipid storage and downregulated expression of lipid metabolism genes. Drosophila STING was found to interact with lipid synthesizing enzymes acetyl-CoA carboxylase (ACC) and fatty acid synthase (FASN). ACC and FASN also interact with each other, indicating that all three proteins may be components of a large multi-enzyme complex. The deletion of Drosophila STING leads to disturbed ACC localization and decreased FASN enzyme activity. Together, these results demonstrate a previously undescribed role of STING in lipid metabolism in Drosophila.
Nagao, K., Yokota, N., Tsuchiya, M., Kato, U., Juni, N., Hara, Y., Mori,
M. X., Mori, Y., Ui-Tei, K., Murate, M., Kobayashi, T., Nishino, Y.,
Miyazawa, A., Yamamoto, A., Suzuki, R., Kaufmann, S., Tanaka, M.,
Tatsumi, K., Nakabe, K., Shintaku, H., Yesylevsky, S., Bogdanov, M. and
Umeda, M. (2021). Extreme deformability of insect cell membranes is governed by phospholipid scrambling. Cell Rep 35(10): 109219. PubMed ID: 34107250
Organization of dynamic cellular structure is crucial for a variety of cellular functions. This study reports that Drosophila and Aedes have highly elastic cell membranes with extremely low membrane tension and high resistance to mechanical stress. In contrast to other eukaryotic cells, phospholipids are symmetrically distributed between the bilayer leaflets of the insect plasma membrane, where phospholipid scramblase (XKR) that disrupts the lipid asymmetry is constitutively active. This study also demonstrates that XKR-facilitated phospholipid scrambling promotes the deformability of cell membranes by regulating both actin cortex dynamics and mechanical properties of the phospholipid bilayer. Moreover, XKR-mediated construction of elastic cell membranes is essential for hemocyte circulation in the Drosophila cardiovascular system. Deformation of mammalian cells is also enhanced by the expression of Aedes XKR.
Friday, September 10th - Behavior
|Behbahani, A. H., Palmer, E. H., Corfas, R. A. and Dickinson, M. H. (2021). Drosophila re-zero their path integrator at the center of a fictive food patch. Curr Biol. PubMed ID: 34450090
The ability to keep track of one's location in space is a critical behavior for animals navigating to and from a salient location, and its computational basis is now beginning to be unraveled. This study tracked flies in a ring-shaped channel as they executed bouts of search triggered by optogenetic activation of sugar receptors. Unlike experiments in open field arenas, which produce highly tortuous search trajectories, the geometrically constrained paradigm enabled monitoring flies' decisions to move toward or away from the fictive food. The results suggest that flies use path integration to remember the location of a food site even after it has disappeared, and flies can remember the location of a former food site even after walking around the arena one or more times. To determine the behavioral algorithms underlying Drosophila search, multiple state transition models were developed and found that flies likely accomplish path integration by combining odometry and compass navigation to keep track of their position relative to the fictive food. The results indicate that whereas flies re-zero their path integrator at food when only one feeding site is present, they adjust their path integrator to a central location between sites when experiencing food at two or more locations. Together, this work provides a simple experimental paradigm and theoretical framework to advance investigations of the neural basis of path integration.
|Vulpe, A., Kim,
H. S., Ballou, S., Wu, S. T., Grabe, V., Nava Gonzales, C., Liang, T.,
Sachse, S., Jeanne, J. M., Su, C. Y. and Menuz, K. (2021). An ammonium transporter is a non-canonical olfactory receptor for ammonia. Curr Biol. PubMed ID: 34111404
Numerous hematophagous insects are attracted to ammonia, a volatile released in human sweat and breath. Low levels of ammonia also attract non-biting insects such as the genetic model organism Drosophila melanogaster and several species of agricultural pests. Two families of ligand-gated ion channels function as olfactory receptors in insects, and studies have linked ammonia sensitivity to a particular olfactory receptor in Drosophila. Given the widespread importance of ammonia to insect behavior, it is surprising that the genomes of most insects lack an ortholog of this gene. This study shows that canonical olfactory receptors are not necessary for responses to ammonia in Drosophila. Instead, it was demonstrated that a member of the ancient electrogenic ammonium transporter family, Amt, is likely a new type of olfactory receptor. This study reports two hitherto unidentified olfactory neuron populations that mediate neuronal and behavioral responses to ammonia in Drosophila. Their endogenous ammonia responses are lost in Amt mutant flies, and ectopic expression of either Drosophila or Anopheles Amt confers ammonia sensitivity. These results suggest that Amt is the first transporter known to function as an olfactory receptor in animals and that its function may be conserved across insect species.
|Stanley, M., Ghosh, B., Weiss, Z. F., Christiaanse, J. and Gordon, M. D. (2021). Mechanisms of lactic acid gustatory attraction in Drosophila. Curr Biol. PubMed ID: 34197729
Sour has been studied almost exclusively as an aversive taste modality. Yet recent work in Drosophila demonstrates that specific carboxylic acids are attractive at ecologically relevant concentrations. This study demonstrates that lactic acid is an appetitive and energetic tastant, which stimulates feeding through activation of sweet gustatory receptor neurons (GRNs). This activation displays distinct, mechanistically separable stimulus onset and removal phases. Ionotropic receptor 25a (IR25a) primarily mediates the onset response, which shows specificity for the lactate anion and drives feeding initiation through proboscis extension. Conversely, sweet gustatory receptors (Gr64a-f) mediate a non-specific removal response to low pH that primarily impacts ingestion. While mutations in either receptor family have marginal impacts on feeding, lactic acid attraction is completely abolished in combined mutants. Thus, specific components of lactic acid are detected through two classes of receptors to activate a single set of sensory neurons in physiologically distinct ways, ultimately leading to robust behavioral attraction.
|Zhang, S. X., Glantz, E. H., Miner, L. E., Rogulja, D. and Crickmore, M. A. (2021). Hormonal control of motivational circuitry orchestrates the transition to sexuality in Drosophila. Sci Adv 7(25). PubMed ID: 34134981
Newborns and hatchlings can perform incredibly sophisticated behaviors, but many animals abstain from sexual activity at the beginning of life. Hormonal changes have long been known to drive both physical and behavioral changes during adolescence, leading to the largely untested assumption that sexuality emerges from organizational changes to neuronal circuitry. This study shows that the transition to sexuality in male Drosophila is controlled by hormonal changes, but this regulation is functional rather than structural. In very young males, a broadly acting hormone directly inhibits the activity of three courtship-motivating circuit elements, ensuring the complete suppression of sexual motivation and behavior. Blocking or overriding these inhibitory mechanisms evokes immediate and robust sexual behavior from very young and otherwise asexual males. Similarities to mammalian adolescence suggest a general principle in which hormonal changes gate the transition to sexuality not by constructing new circuitry but by permitting activity in otherwise latent motivational circuit elements.
|Cho, L. C., Yu, C. C. and Kao, C. F. (2021). Social perception of young adults prolongs the lifespan of aged Drosophila. NPJ Aging Mech Dis 7(1): 21. PubMed ID: 34471134
Lifespan is modulated at distinct levels by multiple factors, including genetic backgrounds, the environment, behavior traits, metabolic status, and more interestingly, sensory perceptions. However, the effects of social perception between individuals living in the same space remain less clear. This study used the Drosophila model to study the influences of social perception on the lifespan of aged fruit flies. The lifespan of aged Drosophila was found to be markedly prolonged after being co-housed with young adults of the same gender. Moreover, the changes of lifespan were affected by several experimental contexts: (1) the ratios of aged and young adults co-housed, (2) the chronological ages of two populations, and (3) the integrity of sensory modalities. Together, it is hypothesize the chemical/physical stimuli derived from the interacting young adults are capable of interfering with the physiology and behavior of aged flies, ultimately leading to the alteration of lifespan.
|Zhu, M. L., Herrera, K. J., Vogt, K. and Bahl, A. (2021). Navigational strategies underlying temporal phototaxis in Drosophila larvae. J Exp Biol 224(11). PubMed ID: 34115116
Navigating across light gradients is essential for survival for many animals. However, there is still a poor understanding of the algorithms that underlie such behaviors. This study developed a novel closed-loop phototaxis assay for Drosophila larvae in which light intensity is always spatially uniform but updates depending on the location of the animal in the arena. Even though larvae can only rely on temporal cues during runs, this study found that they are capable of finding preferred areas of low light intensity. Further detailed analysis of their behavior reveals that larvae turn more frequently and that heading angle changes increase when they experience brightness increments over extended periods of time. It is suggested that temporal integration of brightness change during runs is an important - and so far largely unexplored - element of phototaxis.
Thursday, September 9th
|Neuman, S. D., Lee, A. R., Selegue, J. E., Cavanagh, A. T. and Bashirullah, A. (2021). A novel function for Rab1 and Rab11 during secretory granule maturation. J Cell Sci. PubMed ID: 34224556
Regulated exocytosis is an essential process whereby specific cargo proteins are secreted in a stimulus-dependent manner. Cargo-containing secretory granules are synthesized in the trans-Golgi Network (TGN); after budding from the TGN, granules undergo modifications, including an increase in size. These changes occur during a poorly understood process called secretory granule maturation. This study leveraged the Drosophila larval salivary glands as a model to characterize a novel role for Rab GTPases during granule maturation. Secretory granules were found to increase in size ∼300-fold between biogenesis and release, and loss of Rab1 or Rab11 reduces granule size. Surprisingly, it was found that Rab1 and Rab11 localize to secretory granule membranes. Rab11 associates with granule membranes throughout maturation, and Rab11 recruits Rab1. In turn, Rab1 associates specifically with immature granules and drives granule growth. In addition to roles in granule growth, both Rab1 and Rab11 appear to have additional functions during exocytosis; Rab11 function is necessary for exocytosis, while the presence of Rab1 on immature granules may prevent precocious exocytosis. Overall, these results highlight a new role for Rab GTPases in secretory granule maturation.
|Kuznetsov, I. A. and Kuznetsov, A. V. (2021). Simulation of a sudden drop-off in distal dense core vesicle concentration in Drosophila type II motoneuron terminals. Int J Numer Method Biomed Eng: e3523. PubMed ID: 34418891
Recent experimental observations have shown evidence of an unexpected sudden drop-off in the dense core vesicles (DCVs) content at the ends of certain types of axon endings. A mathematical model was developed that is based on the conservation of captured and transiting DCVs in boutons. The model consists of 77 ordinary differential equations and is solved using a standard Matlab solver. It was hypothesized that the drop in DCV content in distal boutons is due to an insufficient supply of anterogradely moving DCVs coming from the soma. This hypothesis was tested by modifying the flux of DCVs entering the terminal, and it was found that the number of most distal boutons left unfilled increases if the DCV flux entering the terminal is decreased. The number of anterogradely moving DCVs in the axon can be increased either by the release of a portion of captured DCVs into the anterograde component or by an increase of the anterograde DCV flux into the terminal. This increase could lead to having enough anterogradely moving DCVs such that they could reach the most distal bouton and then turn around by changing molecular motors that propel them. The model suggests that this could result in an increased concentration of resident DCVs in distal boutons beginning with bouton 2 (the most distal is bouton 1). This is because in distal boutons, DCVs have a larger chance to be captured from the transiting state as they pass the boutons moving anterogradely and then again as they pass the same boutons moving retrogradely.
|Deruelle, V., Bouillot, S., Job, V., Taillebourg, E., Fauvarque, M. O., Attree, I. and Huber, P. (2021). The bacterial toxin ExoU requires a host trafficking chaperone for transportation and to induce necrosis. Nat Commun 12(1): 4024. PubMed ID: 34188051
Pseudomonas aeruginosa can cause nosocomial infections, especially in ventilated or cystic fibrosis patients. Highly pathogenic isolates express the phospholipase ExoU, an effector of the type III secretion system that acts on plasma membrane lipids, causing membrane rupture and host cell necrosis. This study used a genome-wide screen to discover that ExoU requires DNAJC5, a host chaperone, for its necrotic activity. DNAJC5 is known to participate in an unconventional secretory pathway for misfolded proteins involving anterograde vesicular trafficking. This study shows that DNAJC5-deficient human cells, or Drosophila flies knocked-down for the DNAJC5 orthologue (Cysteine string protein), are largely resistant to ExoU-dependent virulence. ExoU colocalizes with DNAJC5-positive vesicles in the host cytoplasm. DNAJC5 mutations preventing vesicle trafficking (previously identified in adult neuronal ceroid lipofuscinosis, a human congenital disease) inhibit ExoU-dependent cell lysis. These results suggest that, once injected into the host cytoplasm, ExoU docks to DNAJC5-positive secretory vesicles to reach the plasma membrane, where it can exert its phospholipase activity.
|Keith, S. A., Bishop, C., Fallacaro, S. and McCartney, B. M. (2021). Arc1 and the microbiota together modulate growth and metabolic traits in Drosophila. Development. PubMed ID: 34240138
Perturbations to animal-associated microbial communities (the microbiota) have deleterious effects on various aspects of host fitness, but the molecular processes underlying these impacts are poorly understood. This study identified a novel connection between the microbiota and the neuronal factor Arc1 that affects growth and metabolism in Drosophila. Arc1 was found to exhibit tissue-specific microbiota-dependent expression changes, and germ-free flies bearing a null mutation of Arc1 exhibit delayed and stunted larval growth, along with a variety of molecular, cellular, and organismal traits indicative of metabolic dysregulation. Remarkably, it was shown that the majority of these phenotypes can be fully suppressed by mono-association with a single Acetobacter sp. isolate, through mechanisms involving both bacterial diet modification and live bacteria. Additionally, evidence is provided that Arc1 function in key neuroendocrine cells of the larval brain modulates growth and metabolic homeostasis under germ-free conditions. These results reveal a novel role for Arc1 in modulating physiological responses to the microbial environment, and highlight how host-microbe interactions can profoundly impact the phenotypic consequences of genetic mutations in an animal host.
|Du, Q., Chang, J., Cheng, G., Zhao, Y. and Zhou, W. (2021). Sunday Driver Mediates Multi-Compartment Golgi Outposts Defects Induced by Amyloid Precursor Protein. Front Neurosci 15: 673684. PubMed ID: 34140878
Golgi defects including Golgi fragmentation are pathological features of Alzheimer's disease (AD). As a pathogenic factor in AD, amyloid precursor protein (APP) induces Golgi fragmentation in the soma. However, how APP regulates Golgi outposts (GOs) in dendrites remains unclear. Given that APP resides in and affects the movements of GOs, and in particular, reverses the distribution of multi-compartment GOs (mcGOs), this study investigated the regulatory mechanism of mcGO movements in the Drosophila larvae. Knockdown experiments showed that the bidirectional mcGO movements were cooperatively controlled by the dynein heavy chain (Dhc) and kinesin heavy chain subunits. Notably, only Dhc mediated APP's regulation of mcGO movements. Furthermore, by loss-of-function screening, the adaptor protein Sunday driver (Syd) was identified to mediate the APP-induced alteration of the direction of mcGO movements and dendritic defects. Collectively, by elucidating a model of bidirectional mcGO movements, this study revealed the mechanism by which APP regulates the direction of mcGO movements. This study study therefore provides new insights into AD pathogenesis.
|Del Signore, S.
J., Kelley, C. F., Messelaar, E. M., Lemos, T., Marchan, M. F.,
Ermanoska, B., Mund, M., Fai, T. G., Kaksonen, M. and Rodal, A. A.
(2021). An autoinhibitory clamp of actin assembly constrains and directs synaptic endocytosis. Elife 10. PubMed ID: 34324418
Synaptic membrane-remodeling events such as endocytosis require force-generating actin assembly. The endocytic machinery that regulates these actin and membrane dynamics localizes at high concentrations to large areas of the presynaptic membrane, but actin assembly and productive endocytosis are far more restricted in space and time. This study describes a mechanism whereby autoinhibition clamps the presynaptic endocytic machinery to limit actin assembly to discrete functional events. Collective interactions between the Drosophila endocytic proteins Nwk/FCHSD2, Dap160/intersectin, and WASp relieve Nwk autoinhibition and promote robust membrane-coupled actin assembly in vitro. Using automated particle tracking to quantify synaptic actin dynamics in vivo, it was discovered that Nwk-Dap160 interactions constrain spurious assembly of WASp-dependent actin structures. These interactions also promote synaptic endocytosis, suggesting that autoinhibition both clamps and primes the synaptic endocytic machinery, thereby constraining actin assembly to drive productive membrane remodeling in response to physiological cues.
Wednesday, September 8th - Signaling
|Shen, C., Nayak, A., ...., Lee, E., Robbins, D. J. (2021). The E3 ubiquitin ligase component, Cereblon, is an evolutionarily conserved regulator of Wnt signaling. Nat Commun 12(1):5263. PubMed ID: 34489457
Immunomodulatory drugs (IMiDs) are important for the treatment of multiple myeloma and myelodysplastic syndrome. Binding of IMiDs to Cereblon (CRBN), the substrate receptor of the CRL4CRBN E3 ubiquitin ligase, induces cancer cell death by targeting key neo-substrates for degradation. Despite this clinical significance, the physiological regulation of CRBN remains largely unknown. This study demonstrates that Wnt (see Drosophila Wingless), the extracellular ligand of an essential signal transduction pathway, promotes the CRBN-dependent degradation of a subset of proteins. These substrates include Casein kinase 1α (CK1α), a negative regulator of Wnt signaling that functions as a key component of the β-Catenin destruction complex. Wnt stimulation induces the interaction of CRBN with CK1α and its resultant ubiquitination, and in contrast with previous reports does so in the absence of an IMiD. Mechanistically, the destruction complex is critical in maintaining CK1α stability in the absence of Wnt, and in recruiting CRBN to target CK1α for degradation in response to Wnt. CRBN is required for physiological Wnt signaling, as modulation of CRBN in zebrafish and Drosophila yields Wnt-driven phenotypes. These studies demonstrate an IMiD-independent, Wnt-driven mechanism of CRBN regulation and provide a means of controlling Wnt pathway activity by CRBN, with relevance for development and disease.
|Rasmussen, N. R., Smith, H. E. and Reiner, D. J. (2021). The MLK-1/SCD-4 Mixed Lineage Kinase/MAP3K functions to promote dauer formation upstream of DAF-2/InsR. MicroPubl Biol 2021. PubMed ID: 34142023
The C. elegans dauer is an alternative third stage larva induced by dense population and adverse environmental conditions. Genes whose mutants caused dauer formation constitutive (Daf-c) and dauer formation defective (Daf-d) phenotypes were ordered via epistasis into a signaling network, with upstream DAF-7/TGF-beta and DAF-11/receptor guanylyl cyclase defining sensory branches and downstream DAF-2/Insulin receptor and DAF-12/nuclear hormone receptor executing the dauer decision. Mutations in the Scd genes were defined as incompletely penetrant suppressors of the constitutive dauer phenotype conferred by mutation of the DAF-7/TGF-beta signaling axis. SCD-2 was previously shown to be an ortholog of mammalian ALK (Anaplastic Lymphoma Kinase; see Drosophila Alk), a receptor tyrosine kinase. Mutations disrupting the HEN-1/Jeb ligand (see Drosophila Jeb), SOC-1/DOS/GAB adaptor protein and SMA-5/ERK5 atypical MAP Kinase caused Scd phenotypes similar to that of mutant SCD-2. This group regulated expression from a TGF-beta-responsive GFP reporter. This study found that a strain harboring a mutation in the uncharacterized SCD-4 is mutant for MLK-1, the C. elegans ortholog of mammalian Mixed Lineage Kinase and Drosophila slipper (slpr), a MAP3 kinase. This finding was validated by showing that a previously characterized deletion in MLK-1 caused a Scd phenotype similar to that of mutant SCD-4 and altered expression from the TGF-beta-responsive GFP reporter, suggesting that SCD-4 and MLK-1 are the same protein. Based on shared phenotypes and molecular identities, it is hypothesized that MLK-1 functions as a MAP3K in the SCD-2/ALK cascade that signals through SMA-5/ERK5 MAP Kinase to modulate the output of the TGF-beta cascade controlling dauer formation in response to environmental cues.
|Pathak, H. and Varghese, J. (2021). Edem1 activity in the fat body regulates insulin signalling and metabolic homeostasis in Drosophila. Life Sci Alliance 4(8). PubMed ID: 34140347
In Drosophila, nutrient status is sensed by the fat body, a functional homolog of mammalian liver and white adipocytes. The fat body conveys nutrient information to insulin-producing cells through humoral factors which regulate Drosophila insulin-like peptide levels and insulin signalling. Insulin signalling has pleiotropic functions, which include the management of growth and metabolic pathways. This paper reports that Edem1 (endoplasmic reticulum degradation-enhancing α-mannosidase-like protein 1), an endoplasmic reticulum-resident protein involved in protein quality control, acts in the fat body to regulate insulin signalling and thereby the metabolic status in Drosophila Edem1 limits the fat body-derived Drosophila tumor necrosis factor-α Eiger activity on insulin-producing cells and maintains systemic insulin signalling in fed conditions. During food deprivation, edem1 gene expression levels drop, which aids in the reduction of systemic insulin signalling crucial for survival. Overall, this study demonstrates that Edem1 plays a vital role in helping the organism to endure a fluctuating nutrient environment by managing insulin signalling and metabolic homeostasis.
|Yang, G. W. and Tian, Y. (2021). The F-box gene Ppa promotes lipid storage in Drosophila. The F-box gene Ppa promotes lipid storage in Drosophila. Yi Chuan 43(6): 615-622. PubMed ID: 34284991
Lipid is one of the important components of living organisms. The precise regulation and homeostasis maintenance of lipid metabolism are essential to human health. The ubiquitination pathway regulates lipid metabolism by degrading lipid-related proteins. Ppa encodes an F-box protein, which is a member of the SCF ubiquitination complex. Previous studies reported that Ppa regulated the body segmentation and the correct localization of centromere histones, while its function in lipid metabolism has not been reported. In this study, Drosophila melanogaster was used to explore the function of Ppa in lipid storage. The subcellular localization of PPA was detected by fusion with green fluorescent protein. The deletion mutant of Ppa was constructed via CRISPR/Cas9 technology. The morphological changes of lipid droplets in deletion mutants and Ppa overexpression flies were analyzed by BODIPY 493/503 or Nile red staining. Further, Ppa was overexpressed in the deletion mutant to verify its function. The results showed that PPA-GFP fusion protein were localized in the nuclei of salivary gland and fat body. Compared with the control flies, the lipid droplets in Ppa deletion mutants became smaller, and overexpression of Ppa exhibited larger lipid droplets. Overexpression of Ppa in the deletion mutant could restore the lipid droplets to normal state. In summary, this study demonstrated that Ppa could promote lipid storage in Drosophila.
|Aleman, J. R., Kuhn, T. M., Pascual-Garcia, P., Gospocic, J., Lan, Y., Bonasio, R., Little, S. C. and Capelson, M. (2021). Correct dosage of X chromosome transcription is controlled by a nuclear pore component. Cell Rep 35(11): 109236. PubMed ID: 34133927
Dosage compensation in Drosophila melanogaster involves a 2-fold transcriptional upregulation of the male X chromosome, which relies on the X-chromosome-binding males-specific lethal (MSL) complex (see msl-2). However, how such 2-fold precision is accomplished remains unclear. This study shows that a nuclear pore component, Mtor, is involved in setting the correct levels of transcription from the male X chromosome. Using larval tissues, this study demonstrated that the depletion of Mtor results in selective upregulation at MSL targets of the male X, beyond the required 2-fold. Mtor and MSL components interact genetically, and depletion of Mtor can rescue the male lethality phenotype of MSL components. Using RNA fluorescence in situ hybridization (FISH) analysis and nascent transcript sequencing, this study found that the effect of Mtor is not due to defects in mRNA export but occurs at the level of nascent transcription. These findings demonstrate a physiological role for Mtor in the process of dosage compensation, as a transcriptional attenuator of X chromosome gene expression.
|Wang, X. F.,
Yang, S. A., Gong, S., Chang, C. H., Portilla, J. M., Chatterjee, D.,
Irianto, J., Bao, H., Huang, Y. C. and Deng, W. M. (2021). Polyploid mitosis and depolyploidization promote chromosomal instability and tumor progression in a Notch-induced tumor model. Dev Cell 56(13): 1976-1988. PubMed ID: 34146466
Ploidy variation is a cancer hallmark and is frequently associated with poor prognosis in high-grade cancers. Using a Drosophila solid-tumor model where oncogenic Notch drives tumorigenesis in a transition-zone microenvironment in the salivary gland imaginal ring, this study found that the tumor-initiating cells normally undergo endoreplication to become polyploid. Upregulation of Notch signaling, however, induces these polyploid transition-zone cells to re-enter mitosis and undergo tumorigenesis. Growth and progression of the transition-zone tumor are fueled by a combination of polyploid mitosis, endoreplication, and depolyploidization. Both polyploid mitosis and depolyploidization are error prone, resulting in chromosomal copy-number variation and polyaneuploidy. Comparative RNA-seq and epistasis analysis reveal that the DNA-damage response genes, also active during meiosis, are upregulated in these tumors and are required for the ploidy-reduction division. Together, these findings suggest that polyploidy and associated cell-cycle variants are critical for increased tumor-cell heterogeneity and genome instability during cancer progression.
Tuesday, September 7th - Gonads
|Braun, A. L.,
Meghini, F., Villa-Fombuena, G., Guermont, M., Fernandez-Martinez, E.,
Qian, Z., Dolores Martin-Bermudo, M., Gonzalez-Reyes, A., Glover, D. M.
and Kimata, Y. (2021). The careful control of Polo kinase by
APC/C-Ube2C ensures the intercellular transport of germline centrosomes
during Drosophila oogenesis. Open Biol 11(6): 200371. PubMed ID: 34186008
A feature of metazoan reproduction is the elimination of maternal centrosomes from the oocyte. In animals that form syncytial cysts during oogenesis, including Drosophila and human, all centrosomes within the cyst migrate to the oocyte where they are subsequently degenerated. The importance and the underlying mechanism of this event remain unclear. This study shows that, during early Drosophila oogenesis, control of the Anaphase Promoting Complex/Cyclosome (APC/C), the ubiquitin ligase complex essential for cell cycle control, ensures proper transport of centrosomes into the oocyte through the regulation of Polo/Plk1 kinase, a critical regulator of the integrity and activity of the centrosome. This study shows that novel mutations in the APC/C-specific E2, vihar/Ube2c, that affect its inhibitory regulation on APC/C cause precocious Polo degradation and impedes centrosome transport, through destabilization of centrosomes. The failure of centrosome migration correlates with weakened microtubule polarization in the cyst and allows ectopic microtubule nucleation in nurse cells, leading to the loss of oocyte identity. These results suggest a role for centrosome migration in oocyte fate maintenance through the concentration and confinement of microtubule nucleation activity into the oocyte. Considering the conserved roles of APC/C and Polo throughout the animal kingdom, these findings may be translated into other animals.
|Villa-Fombuena, G., Lobo-Pecellin, M., Marin-Menguiano, M., Rojas-Rios, P. and Gonzalez-Reyes, A. (2021). Live
imaging of the Drosophila ovarian niche shows spectrosome and
centrosome dynamics during asymmetric germline stem cell division. Development. PubMed ID: 34370012
Drosophila female germline stem cells (GSCs) are found inside the cellular niche at the tip of the ovary. They undergo asymmetric divisions to renew the stem cell lineage and to produce sibling cystoblasts that will in turn enter differentiation. GSCs and cystoblasts contain spectrosomes, membranous structures essential to orientate the mitotic spindle and that, particularly in GSCs, change shape depending on the cell cycle phase. Using live imaging and a GFP fusion of the spectrosome component Par-1, this study followed the complete spectrosome cycle throughout GSC division and quantified the relative duration of the different spectrosome shapes. It was also determined that the Par-1 kinase shuttles between the spectrosome and the cytoplasm during mitosis, and the continuous addition of new material to the GSC and cystoblast spectrosomes was observed. Next, the Fly-FUCCI tool was used to define in live and fixed tissues that GSCs have a shorter G1 compared to the G2 phase. The observation of centrosomes in dividing GSCs allowed determination that centrosomes separate very early in G1, prior to centriole duplication. Furthermore, this study showed that the anterior centrosome associates with the spectrosome only during mitosis and that, upon mitotic spindle assembly, it translocates to the cell cortex, where it remains anchored until centrosome separation. Finally, this study demonstrated that the asymmetric division of GSCs is not an intrinsic property of these cells, since the spectrosome of GSC-like cells located outside of the niche can divide symmetrically. Thus, GSCs display unique properties during division, a behaviour influenced by the surrounding niche.
|Jang, J. K., Gladstein, A. C., Das, A., Shapiro, J. G., Sisco, Z. L. and McKim, K. S. (2021). Multiple pools of PP2A regulate spindle assembly, kinetochore attachments, and cohesion in Drosophila oocytes. J Cell Sci. PubMed ID: 34160620
Meiosis in female oocytes lacks centrosomes, the microtubule-organizing center. In Drosophila oocytes, meiotic spindle assembly depends on the chromosomal passenger complex (CPC). To investigate the mechanisms that regulate Aurora B activity, the role of Protein Phosphatase 2A (PP2A) in oocyte meiosis was examined. Both forms of PP2A, B55 and B56, antagonize the Aurora B spindle assembly function, suggesting that a balance between Aurora B and PP2A activity maintains the oocyte spindle during meiosis I. PP2A-B56, which is encoded by two partially redundant paralogs, wdb and wrd, is also required for maintaining sister chromatid cohesion, establishing end-on microtubule attachments, and the metaphase I arrest in oocytes. WDB recruitment to the centromeres depends on BUBR1, MEI-S332, and kinetochore protein SPC105R. While BUBR1 stabilizes microtubule attachments in Drosophila oocytes, it is not required for cohesion maintenance during meiosis I. It is proposed that at least three populations of PP2A-B56 regulate meiosis, two of which depend on SPC105R and a third that is associated with the spindle.
|Herbette, M., Wei, X., Chang, C. H., Larracuente, A. M., Loppin, B. and Dubruille, R. (2021). Distinct spermiogenic phenotypes underlie sperm elimination in the Segregation Distorter meiotic drive system. PLoS Genet 17(7): e1009662. PubMed ID: 34228705
Segregation Distorter (SD) is a male meiotic drive system in Drosophila melanogaster. Males heterozygous for a selfish SD chromosome rarely transmit the homologous SD+ chromosome. It is well established that distortion results from an interaction between Sd, the primary distorting locus on the SD chromosome and its target, a satellite DNA called Rsp, on the SD+ chromosome. However, the molecular and cellular mechanisms leading to post-meiotic SD+ sperm elimination remain unclear. This study shows that SD/SD+ males of different genotypes but with similarly strong degrees of distortion have distinct spermiogenic phenotypes. In some genotypes, SD+ spermatids fail to fully incorporate protamines after the removal of histones, and degenerate during the individualization stage of spermiogenesis. In contrast, in other SD/SD+ genotypes, protamine incorporation appears less disturbed, yet spermatid nuclei are abnormally compacted, and mature sperm nuclei are eventually released in the seminal vesicle. This analyses of different SD+ chromosomes suggest that the severity of the spermiogenic defects associates with the copy number of the Rsp satellite. It is proposed that when Rsp copy number is very high (> 2000), spermatid nuclear compaction defects reach a threshold that triggers a checkpoint controlling sperm chromatin quality to eliminate abnormal spermatids during individualization.
|Jensen, L., Venkei, Z. G., Watase, G. J., Bisai, B., Pletcher, S., Lee, C. Y. and Yamashita, Y. M. (2021). me31B regulates stem cell homeostasis by preventing excess dedifferentiation in the Drosophila male germline.
Jensen, L., Venkei, Z. G., Watase, G. J., Bisai, B., Pletcher, S., Lee,
C. Y. and Yamashita, Y. M. (2021). J. Cell Sci. 134(14):jcs258757.
PubMed ID: 34164657
Tissue-specific stem cells maintain tissue homeostasis by providing a continuous supply of differentiated cells throughout the life of organisms. Differentiated/differentiating cells can revert back to a stem cell identity via dedifferentiation to help maintain the stem cell pool beyond the lifetime of individual stem cells. Although dedifferentiation is important for maintaining the stem cell population, it is speculated that it underlies tumorigenesis. Therefore, this process must be tightly controlled. This study shows that a translational regulator, me31B, plays a critical role in preventing excess dedifferentiation in the Drosophila male germline: in the absence of me31B, spermatogonia dedifferentiate into germline stem cells (GSCs) at a dramatically elevated frequency. These results show that the excess dedifferentiation is likely due to misregulation of nos, a key regulator of germ cell identity and GSC maintenance. Taken together, the data reveal negative regulation of dedifferentiation to balance stem cell maintenance with differentiation.
|Neiswender, H., Goldman, C. H., Veeranan-Karmegam, R. and Gonsalvez, G. B. (2021). Dynein light chain-dependent dimerization of Egalitarian is essential for maintaining oocyte fate in Drosophila. Dev Biol 478: 76-88. PubMed ID: 34181915
Egalitarian (Egl) is an RNA adaptor for the Dynein motor and is thought to link numerous, perhaps hundreds, of mRNAs with Dynein. Dynein, in turn, is responsible for the transport and localization of these mRNAs. Studies have shown that efficient mRNA binding by Egl requires the protein to dimerize. It was recently demonstrated that Dynein light chain (Dlc) is responsible for facilitating the dimerization of Egl. Mutations in Egl that fail to interact with Dlc do not dimerize, and as such, are defective for mRNA binding. Consequently, this mutant does not efficiently associate with BicaudalD (BicD), the factor responsible for linking the Egl/mRNA complex with Dynein. This study tested whether artificially dimerizing this Dlc-binding mutant using a leucine zipper would restore mRNA binding and rescue mutant phenotypes in vivo. Interestingly, it was found that although artificial dimerization of Egl restored BicD binding, it only partially restored mRNA binding. As a result, Egl-dependent phenotypes, such as oocyte specification and mRNA localization, were only partially rescued. It was hypothesized that Dlc-mediated dimerization of Egl results in a three-dimensional conformation of the Egl dimer that is best suited for mRNA binding. Although the leucine zipper restores Egl dimerization, it likely does not enable Egl to assemble into the conformation required for maximal mRNA binding activity.
Monday, September 6th - Cell Cycle
Cazorla-Vazquez, S., Ferrazzi, F., Wiederstein, J. L., Grundl, M.,
Weinstock, G., Vergarajauregui, S., Eckstein, M., Kruger, M., Gaubatz,
S. and Engel, F. B. (2021). IQGAP3, a YAP Target, Is Required for Proper Cell-Cycle Progression and Genome Stability. Mol Cancer Res. PubMed ID: 34183451
Controlling cell proliferation is critical for organism development, tissue homeostasis, disease, and regeneration. IQGAP3 (Chd64 in Drosophila) has been shown to be required for proper cell proliferation and migration, and is associated to a number of cancers. Moreover, its expression is inversely correlated with the overall survival rate in the majority of cancers. This study shows that IQGAP3 expression is elevated in cervical cancer and that in these cancers IQGAP3 high expression is correlated with an increased lethality. Furthermore, this study demonstrates that IQGAP3 is a target of YAP (Yorkie in Drosophila), a regulator of cell cycle gene expression. IQGAP3 knockdown resulted in an increased percentage of HeLa cells in S phase, delayed progression through mitosis, and caused multipolar spindle formation and consequentially aneuploidy. Protein-protein interaction studies revealed that IQGAP3 interacts with MMS19, which is known in Drosophila to permit, by competitive binding to Xpd, Cdk7 to be fully active as a Cdk-activating kinase (CAK). Notably, IQGAP3 knockdown caused decreased MMS19 protein levels and XPD knockdown partially rescued the reduced proliferation rate upon IQGAP3 knockdown. This suggests that IQGAP3 modulates the cell cycle via the MMS19/XPD/CAK axis. Thus, in addition to governing proliferation and migration, IQGAP3 is a critical regulator of mitotic progression and genome stability. These data indicate that, while IQGAP3 inhibition might be initially effective in decreasing cancer cell proliferation, this approach harbors the risk to promote aneuploidy and, therefore, the formation of more aggressive cancers.
|Vicars, H., Karg, T., Warecki, B., Bast, I. and Sullivan, W. (2021). Kinetochore-independent mechanisms of sister chromosome separation. PLoS Genet 17(1): e1009304. PubMed ID: 33513180
Although kinetochores normally play a key role in sister chromatid separation and segregation, chromosome fragments lacking kinetochores (acentrics) can in some cases separate and segregate successfully. In Drosophila neuroblasts, acentric chromosomes undergo delayed, but otherwise normal sister separation, revealing the existence of kinetochore- independent mechanisms driving sister chromosome separation. Bulk cohesin removal from the acentric is not delayed, suggesting factors other than cohesin are responsible for the delay in acentric sister separation. In contrast to intact kinetochore-bearing chromosomes, this study discovered that acentrics align parallel as well as perpendicular to the mitotic spindle. In addition, sister acentrics undergo unconventional patterns of separation. For example, rather than the simultaneous separation of sisters, acentrics oriented parallel to the spindle often slide past one another toward opposing poles. To identify the mechanisms driving acentric separation, 117 RNAi gene knockdowns were screened for synthetic lethality with acentric chromosome fragments. In addition to well-established DNA repair and checkpoint mutants, this candidate screen identified synthetic lethality with X-chromosome-derived acentric fragments in knockdowns of Greatwall (cell cycle kinase), EB1 (microtubule plus-end tracking protein), and Map205 (microtubule-stabilizing protein). Additional image-based screening revealed that reductions in Topoisomerase II levels disrupted sister acentric separation. Intriguingly, live imaging revealed that knockdowns of EB1, Map205, and Greatwall preferentially disrupted the sliding mode of sister acentric separation. Based on this analysis of EB1 localization and knockdown phenotypes, it is proposed that in the absence of a kinetochore, microtubule plus-end dynamics provide the force to resolve DNA catenations required for sister separation.
|Shao, B., Diegmiller, R. and Shvartsman, S. Y. (2021). Collective oscillations of coupled cell cycles. Biophys J. PubMed ID: 34197797
Problems with networks of coupled oscillators arise in multiple contexts, commonly leading to the question about the dependence of network dynamics on network structure. Previous work has addressed this question in Drosophila oogenesis, where stable cytoplasmic bridges connect the future oocyte to the supporting nurse cells that supply the oocyte with molecules and organelles needed for its development. To increase their biosynthetic capacity, nurse cells enter the endoreplication program, a special form of the cell cycle formed by the iterated repetition of growth and synthesis phases without mitosis. Recent studies have revealed that the oocyte orchestrates nurse cell endoreplication cycles, based on retrograde (oocyte to nurse cells) transport of a cell cycle inhibitor produced by the nurse cells and localized to the oocyte. Furthermore, the joint dynamics of endocycles has been proposed to depend on the intercellular connectivity within the oocyte-nurse cell cluster. This study used a computational model to argue that this connectivity guides, but does not uniquely determine the collective dynamics and has identified several oscillatory regimes, depending on the time scale of intercellular transport. These results provide insights into collective dynamics of coupled cell cycles and motivate future quantitative studies of intercellular communication in the germline cell clusters.
|Jang, J. K., Gladstein, A. C., Das, A., Shapiro, J. G., Sisco, Z. L. and McKim, K. S. (2021). Multiple pools of PP2A regulate spindle assembly, kinetochore attachments and cohesion in Drosophila oocytes. J Cell Sci 134(14). PubMed ID: 34297127
Meiosis in female oocytes lacks centrosomes, the microtubule-organizing centers. In Drosophila oocytes, meiotic spindle assembly depends on the chromosomal passenger complex (CPC). To investigate the mechanisms that regulate Aurora B activity, this study examined the role of protein phosphatase 2A (PP2A) in Drosophila oocyte meiosis. Both forms of PP2A, B55 and B56, antagonize the Aurora B spindle assembly function, suggesting that a balance between Aurora B and PP2A activity maintains the oocyte spindle during meiosis I. PP2A-B56, which has a B subunit encoded by two partially redundant paralogs, wdb and wrd, is also required for maintenance of sister chromatid cohesion, establishment of end-on microtubule attachments, and metaphase I arrest in oocytes. WDB recruitment to the centromeres depends on BUBR1, MEI-S332 and kinetochore protein SPC105R. Although BUBR1 stabilizes microtubule attachments in Drosophila oocytes, it is not required for cohesion maintenance during meiosis I. At least three populations of PP2A-B56 regulate meiosis are propsed, two of which depend on SPC105R and a third that is associated with the spindle.
|Zhang, P., Katzaroff, A. J., Buttitta, L. A., Ma, Y., Jiang, H., Nickerson, D. W., Ovrebo, J. I. and Edgar, B. A. (2021). The Kruppel-like factor Cabut has cell cycle regulatory properties similar to E2F1. Proc Natl Acad Sci U S A 118(7). PubMed ID: 33558234
Using a gain-of-function screen in Drosophila, the Kruppel-like factor Cabut (Cbt) as a positive regulator of cell cycle gene expression and cell proliferation. Enforced cbt expression is sufficient to induce an extra cell division in the differentiating fly wing or eye, and also promotes intestinal stem cell divisions in the adult gut. Although inappropriate cell proliferation also results from forced expression of the E2f1 transcription factor or its target, Cyclin E, Cbt does not increase E2F1 or Cyclin E activity. Instead, Cbt regulates a large set of E2F1 target genes independently of E2F1, and the data suggest that Cbt acts via distinct binding sites in target gene promoters. Although Cbt was not required for cell proliferation during wing or eye development, Cbt is required for normal intestinal stem cell divisions in the midgut, which expresses E2F1 at relatively low levels. The E2F1-like functions of Cbt identify a distinct mechanism for cell cycle regulation that may be important in certain normal cell cycles, or in cells that cycle inappropriately, such as cancer cells.
|Alvarez-Rodrigo, I., Wainman, A., Saurya, S. and Raff, J. W. (2021). Ana1 helps recruit Polo to centrioles to promote mitotic PCM assembly and centriole elongation. J Cell Sci 134(14). PubMed ID: 34156068
Polo kinase (PLK1 in mammals) is a master cell cycle regulator that is recruited to various subcellular structures, often by its polo-box domain (PBD), which binds to phosphorylated S-pS/pT motifs. Polo/PLK1 kinases have multiple functions at centrioles and centrosomes, and it has been shown that in Drosophila phosphorylated Sas-4 initiates Polo recruitment to newly formed centrioles, while phosphorylated Spd-2 recruits Polo to the pericentriolar material (PCM) that assembles around mother centrioles in mitosis. This study shows that Ana1 (Cep295 in humans) also helps to recruit Polo to mother centrioles in Drosophila. If Ana1-dependent Polo recruitment is impaired, mother centrioles can still duplicate, disengage from their daughters and form functional cilia, but they can no longer efficiently assemble mitotic PCM or elongate during G2. It is concluded that Ana1 helps recruit Polo to mother centrioles to specifically promote mitotic centrosome assembly and centriole elongation in G2, but not centriole duplication, centriole disengagement or cilia assembly (Alvarez-Rodrigo, 2021).
Friday, September 3rd - Disease Models
|Saito, T., Chiku,
T., Oka, M., Wada-Kakuda, S., Nobuhara, M., Oba, T., Shinno, K., Abe,
S., Asada, A., Sumioka, A., Takashima, A., Miyasaka, T. and Ando, K.
(2021). Disulfide bond formation in microtubule-associated tau protein promotes tau accumulation and toxicity in vivo. Hum Mol Genet. PubMed ID: 34137825
Accumulation of microtubule-associated tau protein is thought to cause neuron loss in a group of neurodegenerative diseases called tauopathies. In diseased brains, tau molecules adopt pathological structures that propagate into insoluble forms with disease-specific patterns. Several types of posttranslational modifications in tau are known to modulate its aggregation propensity in vitro, but their influence on tau accumulation and toxicity at the whole-organism level has not been fully elucidated. This study utilized a series of transgenic Drosophila models to compare systematically the toxicity induced by five tau constructs with mutations or deletions associated with aggregation, including substitutions at seven disease-associated phosphorylation sites (S7A and S7E), deletions of PHF6 and PHF6* sequences (ΔPHF6 and ΔPHF6*), and substitutions of cysteine residues in the microtubule binding repeats (C291/322A). Substitutions and deletions resulted in different patterns of neurodegeneration and accumulation, with C291/322A having a dramatic effect on both tau accumulation and neurodegeneration. These cysteines formed disulfide bonds in mouse primary cultured neurons and in the fly retina, and stabilized tau proteins. Additionally, they contributed to tau accumulation under oxidative stress. This study also found that each of these cysteine residues contributes to the microtubule polymerization rate and microtubule levels at equilibrium, but none of them affected tau binding to polymerized microtubules. Since tau proteins expressed in the Drosophila retina are mostly present in the early stages of tau filaments self-assembly, these results suggest that disulfide bond formation by these cysteine residues could be attractive therapeutic targets.
|Putz, S. M., Kram, J., Rauh, E., Kaiser, S., Toews, R., Lueningschroer-Wang, Y., Rieger, D. and Raabe, T. (2021). Loss of p21-activated kinase Mbt/PAK4 causes Parkinson-like phenotypes in Drosophila. Dis Model Mech 14(6). PubMed ID: 34125184
Parkinson's disease (PD) provokes bradykinesia, resting tremor, rigidity and postural instability, and also non-motor symptoms such as depression, anxiety, sleep and cognitive impairments. Similar phenotypes can be induced in Drosophila melanogaster through modification of PD-relevant genes or the administration of PD-inducing toxins. Recent studies correlated deregulation of human p21-activated kinase 4 (PAK4) with PD, leaving open the question of a causative relationship of mutations in this gene for manifestation of PD symptoms. To determine whether flies lacking the PAK4 homolog Mushroom bodies tiny (Mbt) show PD-like phenotypes, a variety of PD criteria was tested. This study demonstrated that mbt mutant flies show PD-like phenotypes including age-dependent movement deficits, reduced life expectancy and fragmented sleep. They also react to a stressful situation with higher immobility, indicating an influence of Mbt on emotional behavior. Loss of Mbt function has a negative effect on the number of dopaminergic protocerebral anterior medial (PAM) neurons, most likely caused by a proliferation defect of neural progenitors. The age-dependent movement deficits are not accompanied by a corresponding further loss of PAM neurons. Previous studies highlighted the importance of a small PAM subgroup for age-dependent PD motor impairments. This study shows that impaired motor skills are caused by a lack of Mbt in this PAM subgroup. In addition, a broader re-expression of Mbt in PAM neurons improves life expectancy. Conversely, selective Mbt knockout in the same cells shortens lifespan. It is concluded that mutations in Mbt/PAK4 can play a causative role in the development of PD phenotypes.
|Wall, J. M., Basu,
A., Zunica, E. R. M., Dubuisson, O. S., Pergola, K., Broussard, J. P.,
Kirwan, J. P., Axelrod, C. L. and Johnson, A. E. (2021). CRISPR/Cas9-engineered Drosophila knock-in models to study VCP diseases. Dis Model Mech 14(7). PubMed ID: 34160014
Mutations in Valosin Containing Protein (VCP) are associated with several degenerative diseases, including multisystem proteinopathy (MSP-1) and amyotrophic lateral sclerosis. However, patients with VCP mutations vary widely in their pathology and clinical penetrance, making it difficult to devise effective treatment strategies. A deeper understanding of how each mutation affects VCP function could enhance the prediction of clinical outcomes and design of personalized treatment options. The power of a genetically tractable model organism coupled with well-established in vivo assays and a relatively short life cycle make Drosophila an attractive system to study VCP disease pathogenesis. Using CRISPR/Cas9, this study generated individual Drosophila knock-in mutants that include nine hereditary VCP disease mutations. The models display many hallmarks of VCP-mediated degeneration, including progressive decline in mobility, protein aggregate accumulation and defects in lysosomal and mitochondrial function. Some novel and unexpected findings were made, including nuclear morphology defects and sex-specific phenotypic differences in several mutants. Taken together, the Drosophila VCP disease models generated in this study will be useful for studying the etiology of individual VCP patient mutations and testing potential genetic and/or pharmacological therapies.
|Xie, J., Chen, S., Bopassa, J. C. and Banerjee, S. (2021). Drosophila tubulin polymerization promoting protein mutants reveal pathological correlates relevant to human Parkinson's disease. Sci Rep. 11(1):13614. PubMed ID: 34193896
Parkinson's disease (PD) is a progressive neurodegenerative disorder with no known cure. PD is characterized by locomotion deficits, nigrostriatal dopaminergic neuronal loss, mitochondrial dysfunctions and formation of α-Synuclein aggregates. A well-conserved and less understood family of Tubulin Polymerization Promoting Proteins (TPPP) is also implicated in PD and related disorders, where TPPP exists in pathological aggregates in neurons in patient brains. However, there are no in vivo studies on mammalian TPPP to understand the genetics and neuropathology linking TPPP aggregation or neurotoxicity to PD. The only Drosophila homolog of human TPPP is named Ringmaker (Ringer). This study reports that adult ringer mutants display progressive locomotor disabilities, reduced lifespan and neurodegeneration. Importantly, the findings reveal that Ringer is associated with mitochondria and ringer mutants have mitochondrial structural damage and dysfunctions. Adult ringer mutants also display progressive loss of dopaminergic neurons. Together, these phenotypes of ringer mutants recapitulate some of the salient features of human PD patients, thus allowing utilization of ringer mutants as a fly model relevant to PD, and further exploration of its genetic and molecular underpinnings to gain insights into the role of human TPPP in PD.
|Wang, L. J., Hsu, T., Lin, H. L. and Fu, C. Y. (2021). Modulation of mitochondrial nucleoid structure during aging and by mtDNA content in Drosophila. Biol Open 10(6). PubMed ID: 34180963
Mitochondrial DNA (mtDNA) encodes gene products that are essential for oxidative phosphorylation. They organize as higher order nucleoid structures (mtNucleoids) that were shown to be critical for the maintenance of mtDNA stability and integrity. While mtNucleoid structures are associated with cellular health, how they change in situ under physiological maturation and aging requires further investigation. This study investigated the mtNucleoid assembly at an ultrastructural level in situ using the TFAM-Apex2 Drosophila model. Smaller and more compact TFAM-nucleoids are populated in the mitochondria of indirect flight muscle of aged flies. Furthermore, mtDNA transcription and replication were cross-regulated in the mtTFB2-knockdown flies as in the mtRNAPol-knockdown flies that resulted in reductions in mtDNA copy numbers and nucleoid-associated TFAM. Overall, this study reveals that the modulation of TFAM-nucleoid structure under physiological aging, which is critically regulated by mtDNA content.
|Ravenscroft, T. A., Phillips, J. B., ..., Westerfield, M. and Bellen, H. J. (2021). Heterozygous loss-of-function variants significantly expand the phenotypes associated with loss of GDF11. Genet Med. PubMed ID: 34113007
Growth differentiation factor 11 (GDF11) is a key signaling protein required for proper development of many organ systems. Only one prior study has associated an inherited GDF11 variant with a dominant human disease in a family with variable craniofacial and vertebral abnormalities. This study expanded the phenotypic spectrum associated with GDF11 variants and documents the nature of the variants. A cohort is presented of six probands with de novo and inherited nonsense/frameshift (4/6 patients) and missense (2/6) variants in GDF11. gdf11 mutant zebrafish were generated to model loss of gdf11 phenotypes, and an overexpression screen was used in Drosophila to test variant functionality. Patients with variants in GDF11 presented with craniofacial (5/6), vertebral (5/6), neurological (6/6), visual (4/6), cardiac (3/6), auditory (3/6), and connective tissue abnormalities (3/6). gdf11 mutant zebrafish show craniofacial abnormalities and body segmentation defects that match some patient phenotypes. Expression of the patients' variants in the fly showed that one nonsense variant in GDF11 is a severe loss-of-function (LOF) allele whereas the missense variants in the cohort are partial LOF variants. GDF11 is needed for human development, particularly neuronal development, and LOF GDF11 alleles can affect the development of numerous organs and tissues.
Thursday, September 2nd - Adult and larval physiology
|Jorgensen, L. B., Malte, H., Orsted, M., Klahn, N. A. and Overgaard, J. (2021). A
unifying model to estimate thermal tolerance limits in ectotherms
across static, dynamic and fluctuating exposures to thermal stress. Sci Rep 11(1): 12840. PubMed ID: 34145337
Temperature tolerance is critical for defining the fundamental niche of ectotherms and researchers classically use either static (exposure to a constant temperature) or dynamic (ramping temperature) assays to assess tolerance. The use of different methods complicates comparison between studies and this study presents a mathematical model (and R-scripts) to reconcile thermal tolerance measures obtained from static and dynamic assays. The model uses input data from several static or dynamic experiments and is based on the well-supported assumption that thermal injury accumulation rate increases exponentially with temperature (known as a thermal death time curve). The model also assumes thermal stress at different temperatures to be additive and using experiments with Drosophila melanogaster, this study validates these central assumptions by demonstrating that heat injury attained at different heat stress intensities and durations is additive. In a separate experiment it was demonstrated that the model can accurately describe injury accumulation during fluctuating temperature stress and further the model was validated by successfully converting literature data of ectotherm heat tolerance (both static and dynamic assays) to a single, comparable metric (the temperature tolerated for 1 h). The model presented in this study has many promising applications for the analysis of ectotherm thermal tolerance and potential pitfalls that should be considered and avoided using this model are discussed.
|Nayak, N. and Mishra, M. (2021). High fat diet induced abnormalities in metabolism, growth, behavior, and circadian clock in Drosophila melanogaster. Life Sci 281: 119758. PubMed ID: 34175317
The current lifestyle trend has made people vulnerable to diabetes and related diseases. Years of scientific research have not been able to yield a cure to the disease completely. The current study aims to investigate a link between high-fat diet mediated diabesity and circadian rhythm in the Drosophila model and inferences that might help in establishing a cure to the dreaded disease. Several experimental methods including phenotypical, histological, biochemical, molecular, and behavioral assays were used in the study to detect obesity, diabetes, and changes in the circadian clock in the fly model. The larva and adults of Drosophila melanogaster exposed to high-fat diet (HFD) displayed excess deposition of fat as lipid droplets and micronuclei formation in the gut, fat body, and crop. Larva and adults of HFD showed behavioral defects. The higher amount of triglyceride, glucose, trehalose in the whole body of larva and adult fly confirmed obesity-induced hyperglycemia. The overexpression of insulin gene (Dilp2) and tribble (trbl) gene expression confirmed insulin resistance in HFD adults. Elevated ROS level, developmental delay, altered metal level, growth defects, locomotory rhythms, sleep fragmentation, and expression of circadian genes (per, tim, and clock) were observed in HFD larva and adults. Thus, HFD impairs the metabolism to produce obesity, insulin resistance, disruption of clock, and circadian clock related co-mordities in D. melanogaster. The circadian gene expression provides an innovative perspective to understand and find a new treatment for type-II diabetes and circadian anomalies.
|Kapila, R., Kashyap, M., Gulati, A., Narasimhan, A., Poddar, S., Mukhopadhaya, A. and Prasad, N. G. (2021). Evolution
of sex-specific heat stress tolerance and larval Hsp70 expression in
populations of Drosophila melanogaster adapted to larval crowding. J Evol Biol. PubMed ID: 34197669
The ability to tolerate temperature stress is an important component of adult fitness. In holometabolous insects like Drosophila melanogaster, adult stress resistance can be affected by growth conditions experienced during the larval stages. Although evolution under crowded larval conditions is known to lead to the correlated evolution of many adult traits, its consequences on adult heat stress tolerance have not been investigated. Therefore, the present study assessed the adult heat stress tolerance in populations of D. melanogaster adapted to a stressful larval crowding environment. Replicate populations of D. melanogaster, selected for adaptation to larval crowding stress (MCUs), were used for more than 230 generations, and their respective controls (MBs). Larvae from selected and control populations were grown under crowded and uncrowded conditions, and their adult heat shock resistance at two different temperatures was measured. Further, Hsp70 expression was compared in crowded and uncrowded larvae of both populations and also measured the Hsp70 expression after a mild heat treatment in adults of selected and control populations. The results showed that adaptation to larval crowding leads to the evolution of Hsp70 gene expression in larval stages and improves adult heat stress tolerance ability in males, but not in females.
|Rodriguez, M., Pagola, L., Norry, F. M. and Ferrero, P. (2021). Cardiac performance in heat-stressed flies of heat-susceptible and heat-resistant Drosophila melanogaster. J Insect Physiol 133: 104268. PubMed ID: 34171365
Thermotolerance is a complex trait that can greatly differ between heat-susceptible (HS) and heat-adapted populations of small insects including Drosophila, with short-term effects after a sub-lethal level of heat stress on many physiological functions. Cardiac performance could accordingly be more robust in heat-resistant (HR) than in HS individuals under heat stress. This study tested heart performance under heat-stress effects in two recombinant inbred lines (RIL) of Drosophila melanogaster that dramatically differ in heat knockdown resistance. Heart rate did not strongly differ between heat-susceptible and heat-tolerant flies after a sub-lethal heat stress. Instead, heat-susceptible flies showed a much higher arrhythmia incidence, a longer duration of each heartbeat, and a larger amount of bradycardia than heat-tolerant flies. The highly conserved cardiac proteins SERCA, RyR and NCX that participate in the excitation/contraction coupling, did not differ in activity level between HR and HS flies. Available information for both RIL suggests that heart performance under heat stress may be linked, at least partially, to candidate genes of previously identified quantitative trait loci (QTL) for thermotolerance. This study indicates that HR flies can be genetically more robust in their heart performance than HS flies under even sub-lethal levels of heat stress.
|Mi, T., Mack, J. O., Lee, C. M. and Zhang, Y. V. (2021). Molecular and cellular basis of acid taste sensation in Drosophila. Nat Commun 12(1): 3730. PubMed ID: 34140480
Acid taste, evoked mainly by protons (H(+)), is a core taste modality for many organisms. The hedonic valence of acid taste is bidirectional: animals prefer slightly but avoid highly acidic foods. However, how animals discriminate low from high acidity remains poorly understood. To explore the taste perception of acid, the fruit fly was used as a model organism. Flies were found to employ two competing taste sensory pathways to detect low and high acidity, and the relative degree of activation of each determines either attractive or aversive responses. Moreover, one member of the fly Otopetrin family, Otopetrin-like a (OtopLa), was established as a proton channel dedicated to the gustatory detection of acid. OtopLa defines a unique subset of gustatory receptor neurons and is selectively required for attractive rather than aversive taste responses. Loss of otopla causes flies to reject normally attractive low-acid foods. Therefore, the identification of OtopLa as a low-acid sensor firmly supports a competition model of acid taste sensation. Altogether, this study has discovered a binary acid-sensing mechanism that may be evolutionarily conserved between insects and mammals.
|Machado, H. E.,
Bergland, A. O., Taylor, R., Tilk, S., Behrman, E., Dyer, K., Fabian,
D. K., Flatt, T., Gonzalez, J., Karasov, T. L., Kim, B., Kozeretska, I.,
Lazzaro, B. P., Merritt, T. J., Pool, J. E., O'Brien, K., Rajpurohit,
S., Roy, P. R., Schaeffer, S. W., Serga, S., Schmidt, P. and Petrov, D.
A. (2021). Broad geographic sampling reveals the shared basis and environmental correlates of seasonal adaptation in Drosophila. eLife. PubMed ID: 34155971
To advance understanding of adaptation to temporally varying selection pressures, signatures of seasonal adaptation occurring in parallel among Drosophila melanogaster populations were identified. Specifically, allele frequencies were assessed genome-wide from flies sampled early and late in the growing season from 20 widely dispersed populations. Parallel seasonal allele frequency shifts were identified across North America and Europe, demonstrating that seasonal adaptation is a general phenomenon of temperate fly populations. Seasonally fluctuating polymorphisms are enriched in large chromosomal inversions, and a broad concordance was found between seasonal and spatial allele frequency change. The direction of allele frequency change at seasonally variable polymorphisms can be predicted by weather conditions in the weeks prior to sampling, linking the environment and the genomic response to selection. These results suggest that fluctuating selection is an important evolutionary force affecting patterns of genetic variation in Drosophila.
Wednesday, September 1st - Stem cells
|Wu, K., Tang, Y., Zhang, Q., Zhuo, Z., Sheng, X., Huang, J., Ye, J., Li, X., Liu, Z. and Chen, H. (2021). Aging-related upregulation of the homeobox gene caudal represses intestinal stem cell differentiation in Drosophila. PLoS Genet 17(7): e1009649. PubMed ID: 34228720
The differentiation efficiency of adult stem cells undergoes a significant decline in aged animals, which is closely related to the decline in organ function and age-associated diseases. However, the underlying mechanisms that ultimately lead to this observed decline of the differentiation efficiency of stem cells remain largely unclear. This study investigated Drosophila midguts and identified an obvious upregulation of caudal (cad), which encodes a homeobox transcription factor. This factor is traditionally known as a central regulator of embryonic anterior-posterior body axis patterning. This study reports that depletion of cad in intestinal stem/progenitor cells promotes quiescent intestinal stem cells (ISCs) to become activate and produce enterocytes in the midgut under normal gut homeostasis conditions. However, overexpression of cad results in the failure of ISC differentiation and intestinal epithelial regeneration after injury. Moreover, this study suggests that cad prevents intestinal stem/progenitor cell differentiation by modulating the Janus kinase/signal transducers and activators of the transcription pathway and Sox21a-GATAe signaling cascade. Importantly, the reduction of cad expression in intestinal stem/progenitor cells restrained age-associated gut hyperplasia in Drosophila. This study identified a function of the homeobox gene cad in the modulation of adult stem cell differentiation and suggested a potential gene target for the treatment of age-related diseases induced by age-related stem cell dysfunction.
|Ray, A., Kamat, K. and Inamdar, M. S. (2021). A
Conserved Role for Asrij/OCIAD1 in Progenitor Differentiation and
Lineage Specification Through Functional Interaction With the Regulators
of Mitochondrial Dynamics. Front Cell Dev Biol 9: 643444. PubMed ID: 34295888
The echanism by which Mitochondria morphology and dynamics regulate cell differentiation and lineage choice remains incompletely understood. Asrij/OCIAD1 is a conserved protein that governs mitochondrial morphology, energy metabolism and human embryonic stem cell (hESC) differentiation. This study compared hESC phenotypes with those of Drosophila hematopoiesis, where Asrij is shown to regulate blood progenitor maintenance by conserved mechanisms. In concordance with hESC studies, this study found that Drosophila Asrij also localizes to mitochondria of larval blood cells and its depletion from progenitors results in elongated mitochondria. Live imaging of asrij knockdown hemocytes and of OCIAD1 knockout hESCs showed reduced mitochondrial dynamics. It was hypothesized that mitochondrial fission and fusion may control progenitor maintenance or differentiation in an Asrij-dependent manner. Knockdown of the fission regulator Drp1 in Drosophila lymph gland progenitors specifically suppressed crystal cell differentiation whereas depletion of the fusion regulator Marf (Drosophila Mitofusin) increased the same with concomitant upregulation of Notch signaling. These phenotypes were stronger in anterior progenitors and were exacerbated by Asrij depletion. Asrij is known to suppress Notch signaling and crystal cell differentiation. This study demonstrates a conserved role for Asrij/OCIAD1 in linking mitochondrial dynamics and progenitor differentiation.
|Riddiford, N., Siudeja, K., van den Beek, M., Boumard, B. and Bardin, A. J. (2021). Evolution and genomic signatures of spontaneous somatic mutation in Drosophila intestinal stem cells. Genome Res. PubMed ID: 34168010
Spontaneous mutations can alter tissue dynamics and lead to cancer initiation. Although large-scale sequencing projects have illuminated processes that influence somatic mutation and subsequent tumor evolution, the mutational dynamics operating in the very early stages of cancer development are currently not well understood. To explore mutational processes in the early stages of cancer evolution, this study exploited neoplasia arising spontaneously in the Drosophila intestine. Analysing whole-genome sequencing data with a dedicated bioinformatic pipeline, this study found neoplasia formation is driven largely through the inactivation of Notch by structural variants, many of which involve highly complex genomic rearrangements. The genome-wide mutational burden in neoplasia was found to be similar to that of several human cancers. Finally, this study identified genomic features associated with spontaneous mutation, and defined the evolutionary dynamics and mutational landscape operating within intestinal neoplasia over the short lifespan of the adult fly. These findings provide unique insight into mutational dynamics operating over a short timescale in the genetic model system, Drosophila melanogaster.
|Sasaki, A., Nishimura, T., Takano, T., Naito, S. and Yoo, S. K. (2021).. White regulates proliferative homeostasis of intestinal stem cells during ageing in Drosophila. Nat Metab 3(4): 546-557. PubMed ID: 33820991
Tissue integrity is contingent on maintaining stem cells. Intestinal stem cells (ISCs) over-proliferate during ageing, leading to tissue dysplasia in Drosophila melanogaster. This study describes a role for white, encoding the evolutionarily conserved ATP-binding cassette transporter subfamily G, with a particularly well-characterized role in eye colour pigmentation, in ageing-induced ISC proliferation in the midgut. ISCs increase expression of white during ageing. ISC-specific inhibition of white suppresses ageing-induced ISC dysregulation and prolongs lifespan. Of the proteins that form heterodimers with White, Brown mediates ISC dysregulation during ageing. Metabolomics analyses reveal previously unappreciated, profound metabolic impacts of white inhibition on organismal metabolism. Among the metabolites affected by White, tetrahydrofolate is transported by White, is accumulated in ISCs during ageing and is indispensable for ageing-induced ISC over-proliferation. Since Thomas Morgan's isolation of a white mutant as the first Drosophila mutant, white mutants have been used extensively as genetic systems and often as controls. These findings provide insights into metabolic regulation of stem cells mediated by the classic gene white.
|Ramesh, P., Dey, N. S., Kanwal, A., Mandal, S. and Mandal, L. (2021). Relish plays a dynamic role in the niche to modulate Drosophila blood progenitor homeostasis in development and infection. Elife 10. PubMed ID: 34292149
Immune challenges demand the gearing up of basal hematopoiesis to combat infection. Little is known about how during development, this switch is achieved to take care of the insult. This study shows that the hematopoietic niche of the larval lymph gland of Drosophila senses immune challenge and reacts to it quickly through the nuclear factor-κB (NF-κB), Relish, a component of the immune deficiency (Imd) pathway. During development, Relish is triggered by ecdysone signaling in the hematopoietic niche to maintain the blood progenitors. Loss of Relish causes an alteration in the cytoskeletal architecture of the niche cells in a Jun Kinase dependent manner, resulting in the trapping of Hh implicated in progenitor maintenance. Notably, during infection, downregulation of Relish in the niche tilts the maintenance program towards precocious differentiation, thereby bolstering the cellular arm of the immune response.
|Resnik-Docampo, M., Cunningham, K. M., Ruvalcaba, S. M., Choi, C., Sauer, V. and Jones, D. L. (2021). Neuroglian
regulates Drosophila intestinal stem cell proliferation through
enhanced signaling via the epidermal growth factor receptor. Stem Cell Reports. PubMed ID: 33961791
The Drosophila intestine is an excellent system for elucidating mechanisms regulating stem cell behavior. This study shows that the septate junction (SJ) protein Neuroglian (Nrg) is expressed in intestinal stem cells (ISCs) and enteroblasts (EBs) within the fly intestine. SJs are not present between ISCs and EBs, suggesting Nrg plays a different role in this tissue. This study reveals that Nrg is required for ISC proliferation in young flies, and depletion of Nrg from ISCs and EBs suppresses increased ISC proliferation in aged flies. Conversely, overexpression of Nrg in ISC and EBs promotes ISC proliferation, leading to an increase in cells expressing ISC/EB markers; in addition, an increase was observed in epidermal growth factor receptor (Egfr) activation. Genetic epistasis experiments reveal that Nrg acts upstream of Egfr to regulate ISC proliferation. As Nrg function is highly conserved in mammalian systems, this work characterizing the role of Nrg in the intestine has implications for the treatment of intestinal disorders that arise due to altered ISC behavior.
|Ghotbi, E., Ye, P., Ervin, T., Kum, A., Benes, J. and Jones, R. S. (2021). Polycomb-group recruitment to a Drosophila target gene is the default state that is inhibited by a transcriptional activator. Sci Adv 7(29). PubMed ID: 34272248
Polycomb-group (PcG) proteins are epigenetic regulators that maintain the transcriptional repression of target genes following their initial repression by transcription factors. PcG target genes are repressed in some cells, but active in others. Therefore, a mechanism must exist by which PcG proteins distinguish between the repressed and active states and only assemble repressive chromatin environments at target genes that are repressed. This study presents experimental evidence that the repressed state of a Drosophila PcG target gene, giant (gt), is not identified by the presence of a repressor. Rather, de novo establishment of PcG-mediated silencing at gt is the default state that is prevented by the presence of an activator or coactivator, which may inhibit the catalytic activity of Polycomb-repressive complex 2 (PRC2).
|Bag, I., Chen, S., Rosin, L. F., Chen, Y., Liu, C. Y., Yu, G. Y. and Lei, E. P. (2021). M1BP cooperates with CP190 to activate transcription at TAD borders and promote chromatin insulator activity. Nat Commun 12(1): 4170. PubMed ID: 34234130
Genome organization is driven by forces affecting transcriptional state, but the relationship between transcription and genome architecture remains unclear. This study identified the Drosophila transcription factor Motif 1 Binding Protein (M1BP) in physical association with the gypsy chromatin insulator core complex, including the universal insulator protein CP190. M1BP is required for enhancer-blocking and barrier activities of the gypsy insulator as well as its proper nuclear localization. Genome-wide, M1BP specifically colocalizes with CP190 at Motif 1-containing promoters, which are enriched at topologically associating domain (TAD) borders. M1BP facilitates CP190 chromatin binding at many shared sites and vice versa. Both factors promote Motif 1-dependent gene expression and transcription near TAD borders genome-wide. Finally, loss of M1BP reduces chromatin accessibility and increases both inter- and intra-TAD local genome compaction. These results reveal physical and functional interaction between CP190 and M1BP to activate transcription at TAD borders and mediate chromatin insulator-dependent genome organization.
Monday, August 2nd - Cytoskeleton and Junctions
|Chung, S., Le, T. P., Vishwakarma, V., Cheng, Y. L. and Andrew, D. J. (2021). Isoform-specific roles of the Drosophila filamin-type protein Jitterbug (Jbug) during development. Genetics. PubMed ID: 34173831
Filamins are highly conserved actin-crosslinking proteins that regulate organization of the actin cytoskeleton. As key components of versatile signaling scaffolds, filamins are implicated in developmental anomalies and cancer. Multiple isoforms of filamins exist, raising the possibility of distinct functions for each isoform during development and in disease. This study provides an initial characterization of jitterbug (jbug), which encodes one of the two filamin-type proteins in Drosophila. Jbug antiserum was generated that recognizes all of the spliced forms and reveals differential expression of different Jbug isoforms during development, and a significant maternal contribution of Jbug protein. To reveal the function of Jbug isoforms, new genetic tools were developed, including a null allele that deletes all isoforms, hypomorphic alleles that affect only a subset, and UAS lines for Gal4-driven expression of the major isoforms. Using these tools, it was demonstrated that Jbug is required for viability and that specific isoforms are required in the formation of actin-rich protrusions including thoracic bristles in adults and ventral denticles in the embryo. Specific isoforms of Jbug show differential localization within epithelia, and maternal and zygotic loss of jbug disrupts Crumbs (Crb) localization in several epithelial cell types.
|Herrera-Perez, R. M., Cupo, C., Allan, C., Lin, A. and Kasza, K. E. (2021). Using optogenetics to link myosin patterns to contractile cell behaviors during convergent extension. Biophys J. PubMed ID: 34293302
Distinct patterns of actomyosin contractility are often associated with particular epithelial tissue shape changes during development. For example, a planar polarized pattern of myosin II localization regulated by Rho1 signaling during Drosophila body axis elongation is thought to drive cell behaviors that contribute to convergent extension. This study developed optogenetic tools to activate (optoGEF) or deactivate (optoGAP) Rho1 signaling. These tools were used to manipulate myosin patterns at the apical side of the germband epithelium during Drosophila axis elongation, and the effects on contractile cell behaviors were analyzed. Uniform activation or inactivation of Rho1 signaling across the apical surface of the germband is sufficient to disrupt the planar polarized pattern of myosin at cell junctions on the timescale of 3-5 min, leading to distinct changes in junctional and medial myosin patterns in optoGEF and optoGAP embryos. These two perturbations to Rho1 activity both disrupt axis elongation and cell intercalation These studies demonstrate that acute optogenetic perturbations to Rho1 activity are sufficient to rapidly override the endogenous planar polarized myosin pattern in the germband during axis elongation. Moreover, these results reveal that the levels of Rho1 activity and the balance between medial and junctional myosin play key roles, not only in organizing the cell rearrangements that are known to directly contribute to axis elongation, but also in regulating cell area fluctuations and cell packings, which have been proposed to be important factors influencing the mechanics of tissue deformation and flow.
|Bonello, T., Aguilar-Aragon, M., Tournier, A. and Thompson, B. J. (2021). A picket fence function for adherens junctions in epithelial cell polarity. Cells Dev: 203719. PubMed ID: 34242843
Adherens junctions are a defining feature of all epithelial cells, providing cell-cell adhesion and contractile ring formation that is essential for cell and tissue morphology. In Drosophila, adherens junctions are concentrated between the apical and basolateral plasma membrane domains, defined by aPKC-Par6-Baz and Lgl/Dlg/Scrib, respectively. Whether adherens junctions contribute to apical-basal polarization itself has been unclear because neuroblasts exhibit apical-basal polarization of aPKC-Par6-Baz and Lgl in the absence of adherens junctions. This study shows that, upon disruption of adherens junctions in epithelial cells, apical polarity determinants such as aPKC can still segregate from basolateral Lgl, but lose their sharp boundaries and also overlap with Dlg and Scrib - similar to neuroblasts. In addition, control of apical versus basolateral domain size is lost, along with control of cell shape, in the absence of adherens junctions. Manipulating the levels of apical Par3/Baz or basolateral Lgl polarity determinants in experiments and in computer simulations confirms that adherens junctions provide a 'picket fence' diffusion barrier that restricts the spread of polarity determinants along the membrane to enable precise domain size control. Movement of adherens junctions in response to mechanical forces during morphogenetic change thus enables spontaneous adjustment of apical versus basolateral domain size as an emergent property of the polarising system.
|Gillard, G., Girdler, G. and Roper, K. (2021). A release-and-capture mechanism generates an essential non-centrosomal microtubule array during tube budding. Nat Commun 12(1): 4096. PubMed ID: 34215746
Non-centrosomal microtubule arrays serve crucial functions in cells, yet the mechanisms of their generation are poorly understood. During budding of the epithelial tubes of the salivary glands in the Drosophila embryo, it has been demonstrated that the activity of pulsatile apical-medial actomyosin depends on a longitudinal non-centrosomal microtubule array. This study uncovered that the exit from the last embryonic division cycle of the epidermal cells of the salivary gland placode leads to one centrosome in the cells losing all microtubule-nucleation capacity. This restriction of nucleation activity to the second, Centrobin-enriched, centrosome is key for proper morphogenesis. Furthermore, the microtubule-severing protein Katanin and the minus-end-binding protein Patronin accumulate in an apical-medial position only in placodal cells. Loss of either in the placode prevents formation of the longitudinal microtubule array and leads to loss of apical-medial actomyosin and impaired apical constriction. A mechanism is proposed whereby Katanin-severing at the single active centrosome releases microtubule minus-ends that are then anchored by apical-medial Patronin to promote formation of the longitudinal microtubule array crucial for apical constriction and tube formation.
|Nabais, C., Pessoa,
D., de-Carvalho, J., van Zanten, T., Duarte, P., Mayor, S., Carneiro,
J., Telley, I. A. and Bettencourt-Dias, M. (2021). Plk4 triggers autonomous de novo centriole biogenesis and maturation. J Cell Biol 220(5). PubMed ID: 33760919
Centrioles form centrosomes and cilia. In most proliferating cells, centrioles assemble through canonical duplication, which is spatially, temporally, and numerically regulated by the cell cycle and the presence of mature centrioles. However, in certain cell types, centrioles assemble de novo, yet by poorly understood mechanisms. This study established a controlled system to investigate de novo centriole biogenesis, using Drosophila melanogaster egg explants overexpressing Polo-like kinase 4 (Plk4), a trigger for centriole biogenesis. At a high Plk4 concentration, centrioles form de novo, mature, and duplicate, independently of cell cycle progression and of the presence of other centrioles. Plk4 concentration determines the temporal onset of centriole assembly. Moreover, the results suggest that distinct biochemical kinetics regulate de novo and canonical biogenesis. Finally, which other factors modulate de novo centriole assembly was investigate, and proteins of the pericentriolar material (PCM), and in particular γ-tubulin, were found to promote biogenesis, likely by locally concentrating critical components.
|Hahn, I., Voelzmann, A., Parkin, J., Fulle, J. B., Slater, P. G., Lowery, L. A., Sanchez-Soriano, N. and Prokop, A. (2021). Tau, XMAP215/Msps and Eb1 co-operate interdependently to regulate microtubule polymerisation and bundle formation in axons. PLoS Genet 17(7): e1009647. PubMed ID: 34228717
The formation and maintenance of microtubules requires their polymerisation, but little is known about how this polymerisation is regulated in cells. Focussing on the essential microtubule bundles in axons of Drosophila and Xenopus neurons, this study showed that the plus-end scaffold Eb1, the polymerase XMAP215/Msps and the lattice-binder Tau co-operate interdependently to promote microtubule polymerisation and bundle organisation during axon development and maintenance. Eb1 and XMAP215/Msps promote each other's localisation at polymerising microtubule plus-ends. Tau outcompetes Eb1-binding along microtubule lattices, thus preventing depletion of Eb1 tip pools. The three factors genetically interact and show shared mutant phenotypes: reductions in axon growth, comet sizes, comet numbers and comet velocities, as well as prominent deterioration of parallel microtubule bundles into disorganised curled conformations. This microtubule curling is caused by Eb1 plus-end depletion which impairs spectraplakin-mediated guidance of extending microtubules into parallel bundles. This demonstration that Eb1, XMAP215/Msps and Tau co-operate during the regulation of microtubule polymerisation and bundle organisation, offers new conceptual explanations for developmental and degenerative axon pathologies.
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