Sox102F: Biological Overview | References
Gene name - Sox102F
Synonyms - SoxD
Cytological map position -
Function - transcription factor
Symbol - Sox102F
FlyBase ID: FBgn0039938
Genetic map position - chr4:800,436-804,080
NCBI classification - SOX-TCF_HMG-box
Cellular location - cytoplasmic
|Recent literature||Schilling, T., Ali, A. H., Leonhardt, A., Borst, A. and Pujol-Marti, J. (2019). Transcriptional control of morphological properties of direction-selective T4/T5 neurons in Drosophila. Development. PubMed ID: 30642835
In the Drosophila visual system, T4/T5 neurons represent the first stage in which the direction of visual motion is computed. T4 and T5 neurons exist in four subtypes, each responding to motion in one of the four cardinal directions and projecting axons into one of the four lobula plate layers. However, all T4/T5 neurons share properties essential for sensing motion. How T4/T5 neurons acquire their properties during development is poorly understood. This study reveals that SoxN and Sox102F transcription factors control the acquisition of properties common to all T4/T5 neuron subtypes, i.e. the layer specificity of dendrites and axons. Accordingly, adult flies are motion blind after disrupting SoxN or Sox102F in maturing T4/T5 neurons. It was further foud that the transcription factors Ato and Dac are redundantly required in T4/T5 neuron progenitors for SoxN and Sox102F expression in T4/T5 neurons, linking the transcriptional programs specifying progenitor identity to those regulating the acquisition of morphological properties in neurons. This work will help to link structure, function and development in a neuronal type performing a computation conserved across vertebrate and invertebrate visual systems.
Precise control of neurite guidance during development is essential to ensure proper formation of neuronal networks and correct function of the central nervous system (CNS). How neuronal projections find their targets to generate appropriate synapses is not entirely understood. Although transcription factors are key molecules during neurogenesis, their entire function during the formation of networks in the CNS is not known. This study used the Drosophila melanogaster optic lobe as a model for understanding neurite guidance during development. The function of Sox102F/SoxD, the unique Drosophila orthologue of the vertebrate SoxD family of transcription factors, was assessed. SoxD is expressed in immature and mature neurons in the larval and adult lobula plate ganglia (one of the optic lobe neuropils), but is absent from glial cells, neural stem cells and progenitors of the lobula plate. SoxD RNAi knockdown in all neurons results in a reduction of the lobula plate neuropil, without affecting neuronal fate. This morphological defect is associated with an impaired optomotor response of adult flies. Moreover, knocking down SoxD only in T4/T5 neuronal types, which control motion vision, affects proper neurite guidance into the medulla and lobula. These findings suggest that SoxD regulates neurite guidance, without affecting neuronal fate (Contreras, 2018).
The Drosophila melanogaster visual system is composed of the retina and the optic lobe, which is divided into four ganglia: lamina, medulla, lobula and lobula plate. The visual inputs travel from the retinal photoreceptors through different optic lobe neurons, where this information is processed, triggering behavioural responses. Correct connectivity between optic lobe neurons is fundamental for sensing visual information. In the past few years, several studies have characterised how transcription factors regulate the development and neuronal composition of the optic lobe. However, while the development of the lamina and medulla has been extensively studied, research has only recently been focused on the development of the lobula complex (lobula and lobula plate) (Contreras, 2018).
The Sox (Sry Box) family of transcription factors is a key regulator of embryonic development. These transcription factors bear a conserved DNA binding domain known as the SRY-related High Mobility Group-box (HMG-box), which was first described in the Sry protein that is fundamental for sex determination in mammals. The Sox family of proteins is subdivided into nine groups, depending on the amino acid composition of their HMG-box. Vertebrate genomes encode approximately 20 members of the Sox family, whereas only eight members have been described in the fruit fly. The Sox transcription factors do not activate or repress gene expression themselves, but act together with partner factors that determine the modulation of target genes (Contreras, 2018).
Sox proteins are important during neural development and different groups of Sox transcription factors are responsible for similar neurodevelopmental processes across species. For instance, SoxB1 group members work in early neurogenesis in vertebrates and invertebrates. The vertebrate SoxB1 protein, Sox2, participates in early events of central and peripheral nervous system development. In a similar manner, the SoxB1 orthologues in Drosophila, SoxNeuro (SoxN) and Dichaete, are also required for proper neurogenesis and the formation of neural stem cells during development. On the other hand, SoxD proteins are generally involved later during nervous system development. Vertebrate Sox5 and Sox6 regulate neural stem cell proliferation, neuronal diversity, neuronal migration and projection formation. Similarly, the Drosophila SoxD orthologue is necessary for the development of the nervous system and loss of SoxD function affects synaptic bouton development at the neuromuscular junction and dendritic arborisation in sensory neurons (Contreras, 2018).
This study analysed the role of the Drosophila melanogaster orthologue of the SoxD family: Sox102F/SoxD during optic lobe development. SoxD is expressed in all optic lobe ganglia and is involved in the morphogenesis of the lobula plate neuropil. RNAi-mediated SoxD knockdown in developing neurons severely alters the morphology of the lobula plate ganglia. These morphological defects are not a consequence of changes in the fate of lobula plate neurons, but result from an alteration in the normal pattern of axon and dendrite formation. Associated with the defects in lobula plate morphology, the fly optomotor response is also impaired upon SoxD downregulation in lobula plate neurons. These results are consistent with the observation that Sox5 is involved in neuronal migration and axon pathfinding in mice, denoting that SoxD function is evolutionary conserved (Contreras, 2018).
According to a phylogenetic analysis, Sox102F has high homology to human Sox5, Sox6 and Sox13 transcription factors; therefore, it is proposed to rename this protein SoxD. During larval brain development, SoxD is expressed in neurons of the medulla and the lobula complex, while in the lamina, SoxD is transiently expressed before LPC differentiation into neurons. SoxD knockdown in all neurons or in lobula plate neurons severely affects the morphology of the lobula plate neuropil, impairing fly optomotor behaviour. Thus, this study shows that SoxD is required for the control of lobula plate T4 and T5 neurite guidance without affecting neuronal fate. Finally, it was demonstrated that misexpression of SoxD is sufficient to alter neurite guidance in photoreceptors and mushroom body Kenyon cells (Contreras, 2018).
The lobula plate is one of the less explored ganglia of the Drosophila visual system. Although neurogenesis has been described in some detail, later stages of development and the mechanisms directing neuronal subtype specification are starting to be described (Pinto-Teixeira, 2018). The lobula plate has an important role in motion detection in insects, while the lobula has a role in the integration of optic stimuli and the behavioural response. Two important neuronal populations in this regard are T4 and T5 neurons, which gather information from the medulla and lobula respectively and make their synaptic outputs in the lobula plate. Upon SoxD knockdown in all neurons (Elav-GAL4), specifically in T4/T5 neurons (IPC-GAL4 plus R42F06-Gal4) or only in T5 neurons (R42H07-GAL4), defects are observed in lobula plate morphology with increasing severity correlated with the number of affected neurons. This result suggests that SoxD may be also required for the developing of other lobula plate neurons and not only for T4/T5 neurons. In addition to RNAi-mediated knockdown experiments, a combination of soxD alleles was used that reduces the gene dosage. This hypomorphic condition showed a similar but milder phenotype, supporting the specificity of this phenotype to SoxD function. Furthermore, analysing SoxD knockdown phenotype in T4 or T5 neurons, alterations were observed in axon/dendrite guidance that could explain the motion perception defects of the adult animal (Contreras, 2018).
Recently, Li (2017) showed that SoxD is involved in neuronal development and degeneration. The Li study demonstrated that SoxD is required for synaptic bouton development at the neuromuscular junction, dendritic arborisation in sensory neurons, olfactory behaviour and climbing. This study describes a similar role for SoxD in the development of the lobula plate, but no evidence was found of apoptosis upon SoxD knockdown in larval stages, suggesting that the phenotype observed was not due to loss of lobula plate neurons but to neurite mistargeting during development. Moreover, it wasshowm that optomotor behaviour is also affected after SoxD knockdown, complementing Li's behavioural observations. This evidence lends support for the role of SoxD in axon and dendrite guidance in different types of neurons of the CNS (Contreras, 2018).
The mechanisms by which lobula plate neurite guidance is controlled by SoxD are unknown. Recently, it was described that Atonal promotes the differentiation of T4 and T5 neurons, while Notch signalling activity discriminates between T4 and T5 neuronal fates. This study observed that loss of SoxD did not affect T4/T5 differentiation markers, suggesting that the function of SoxD lays downstream of the T4/T5 fate decision. Thus, it is proposed that SoxD controls final stages of neuronal differentiation during development (Contreras, 2018).
The overgrowth of neurites observed upon SoxD knockdown in T4 and T5 neurons may result from defects in sensing an inhibitory guidance cue that restrict their growth into the medulla or the lobula. In accordance to this hypothesis, SoxD misexpression strongly affects neurite guidance in at least two different systems: photoreceptors and Kenyon cells. This supports the possible role of SoxD on sensing guidance cues. Additionally, the presence of the synaptic terminal marker in T5 dendrites after SoxD knockdown, suggests problems in neurite differentiation that could contribute to the guidance defects. Interestingly, the R42H07-GAL4 driver used to target T5 neurons was generated using an enhancer from the soxD locus. This enhancer does not recapitulate the entire expression of soxD, which is also expressed in T4 neurons and other lobula plate neurons. However, SoxD knockdown was associated to an increase of the GFP fluorescence driven by R42H07-GAL4, suggesting that SoxD may negatively regulate this enhancer (Contreras, 2018).
The regulation of neurite guidance may not be the only role of Drosophila SoxD during development. Over-expression of SoxD in embryonic neuroblasts and RNAi-mediated knockdown of SoxD in glial cells were reported to severely disrupt embryonic CNS development. Surprisingly, this study did not observe expression of SoxD in glial cells in the larval brain, while it remains unknown whether SoxD is expressed in embryonic glia. Furthermore, SoxD is also relevant for the function of other organs. SoxD knockdown in cardiac cells affects heart anatomy and function, while SoxD-RNAi expressed in wing discs increased the size of the longitudinal veins L2 and L3, and the marginal vein (Contreras, 2018).
Future work should address the signalling pathways upstream of SoxD activation and the SoxD targets that govern the morphogenesis of lobula plate neurons. Interestingly, Sox5 knockout mice show defects in axonal pathfinding of corticothalamic neurons, similar to Drosophila T4/T5 neurons, suggesting a conserved role of SoxD proteins in neurite guidance (Contreras, 2018).
A recent paper depicted the role of Sox5 in the regulation of the Collapsin Response Mediator Protein (CRMP), an intracellular protein involved in neurite guidance (Naudet, 2018). Interestingly the authors showed that Sox5 gain of function reduces neurite guidance through CRMP in hippocampal neurons, shedding some light to the molecular mechanism involved. Future work should address the conservation of this regulation (Contreras, 2018).
Finally, human Sox5 has been implicated in a number of diseases and intellectual disability in humans. Several studies report mutations and deletions in the sox5 locus that are linked to developmental defects. Therefore, using the fly as a model for neurite guidance may be valuable in determining the biological impact of these mutations in the onset of neurological diseases (Contreras, 2018).
SOX5 encodes a transcription factor that is expressed in multiple tissues including heart, lung and brain. Mutations in SOX5 have been previously found in patients with amyotrophic lateral sclerosis (ALS) and developmental delay, intellectual disability and dysmorphic features. To characterize the neuronal role of SOX5, this study silenced the Drosophila ortholog of SOX5, Sox102F, by RNAi in various neuronal subtypes in Drosophila. Silencing of Sox102F leads to misorientated and disorganized microchaetes, neurons with shorter dendritic arborization (DA) and reduced complexity, diminished larval peristaltic contractions, loss of neuromuscular junction bouton structures, impaired olfactory perception, and severe neurodegeneration in brain. Silencing of SOX5 in human SH-SY5Y neuroblastoma cells results in a significant repression of WNT signaling activity and altered expression of WNT-related genes. Samples of SOX5 variants reveals several variants that show significant association with Alzheimers disease disease status. These findings indicate that SOX5 is a novel candidate gene for AD with important role in neuronal function. The genetic findings warrant further studies to identify and characterize SOX5 variants that confer risk for AD, ALS and intellectual disability (Li, 2017).
This comprehensive study of SOX5 was carried out in transgenic Drosophila models, human SH-SY5Y neuroblastoma cells and LOAD families, respectively. Silencing of Sox102F in Drosophila leads to destructive neuron and abnormal behavior and SOX5 regulates the WNT signaling pathway activity and the expression of the WNT-related genes in human neuronal cells. It was also observed that variants in SOX5 were associated with AD by family-based GWAS, and novel mutations in AD families using whole genome sequence analyses (Li, 2017).
Studies have shown that SOX5 modulates cell fate, controls cell proliferation, regulates cartilage formation and neuronal development. In mouse, Sox5 plays important roles in the specification of subcortically projecting axons in the developing cerebral cortex. In Sox5-null mice, the Sox5-null neocortex neurons maintain an immature differentiation state by losing and misrouting axons; Sox5 overexpression causes re-emergence of neurons with corticofugal features. Sox5 is expressed in the chick neural crest and neural progenitors and is required for the development of the glial lineage and neuronal differentiation. Overall, previous studies indicate an important role for SOX5 in neuronal generation, differentiation and diversity (Li, 2017).
To characterize the functional role of SOX5 in adult neuron and brain function, Sox102F, the Drosophila ortholog of SOX5, was silenced in various neuronal subtypes. Silencing of Sox102F led to misorientated and disorganized michrochaetes, shorter DA neurons with lower complexity, reduced contraction movements and loss of neuron muscular junction bouton structure, impaired olfactory perception and severe neuron degeneration in whole brain. These novel findings implicate an important functional role of SOX5 in neuronal development and brain function (Li, 2017).
The cognitive hallmark of AD is an extraordinary inability to form new memories. Precise and rapid synaptic vesicle recycling is crucial for synaptic transmission and plays an important role in synaptic plasticity. At the NMJ, pre-synaptic motor neurons depolarize the post-synaptic muscle. Silencing of Sox102F resulted in loss of NMJ bouton structure. In addition, MBs are the learning and memory center of Drosophila, silencing of Sox102F led to an impaired olfactory perception. Furthermore, courtship behavior has been widely used to examine the activity and coordination, as well as learning and memory, as it involves the exchange of various sensory stimuli including visual, auditory, and chemosensory signals between males and females, which lead to a complex series of well characterized motor behaviors culminating in successful copulation. Silencing of Sox102F significantly affected adult courtship behavior and exhibited a significantly delay in the mating steps (licking and curling), a statistically significant shortened length of copulation and a reduced CI. Moreover, silencing of Sox102F in whole brain led to severe neurodegeneration. Histological analysis in adult brain showed the degenerated brain structure, which was consistent with behavioral changes (Li, 2017).
The WNT family of secreted growth factors regulates the developmental processes of cell fate and polarity and cell maintenance processes. WNT signaling consists of canonical WNT signaling (WNT/β-catenin pathway) that is dependent on the β-catenin pathway, whereas noncanonical WNT signaling involves β-catenin independent pathways, which comprise different types of WNT ligands and receptors. Aβ binds to the extracellular cysteine-rich domain of the Frizzled receptor (Fz) inhibiting WNT/β-catenin signaling. Activation of WNT signaling by Huperzine A enhances the nonamyloidogenic pathway in an AD transgenic mouse model, suggesting a sustained loss of WNT signaling function may be involved in the Aβ-dependent neurodegeneration observed in AD brain. Sox5 regulates the timing of cell cycle exit by triggering the expression of the negative regulator of the pathway axin2, which opposes WNT-beta-catenin activity on cell cycle progression in chick. In addition, previous work has found that silencing of Sox102F increased the WNT-related Wg expression in the wing disc of Drosophila (Li, 2013). These previous studies supported a functional role for SOX5 in WNT signaling. This study now demonstrates that silencing of SOX5 resulted in a statistically significant repression of WNT signaling pathway activity. SOX5 regulated the expression of 7 of the 84 genes related to WNT signaling pathway including GSK3β. GSK3β is a constitutively active, proline-directed serine/threonine kinase that directly controls the level of β-catenin phosphorylation, which leads to its consequent degradation by the proteasome pathway. Studies have shown that GSK3β plays a pivotal and central role in the pathogenesis of both sporadic and familial forms of AD. GSK3β is involved in the formation of paired helical filament (PHF)-tau, which is an integral component of the neurofibrillary tangle (NFT) deposits that disrupt neuronal function. It is suggested that over-activity of GSK3β accounts for memory impairment, tau hyper-phosphorylation, increased β-amyloid production and local plaque-associated microglial-mediated inflammatory responses; all of which are hallmark characteristics of AD. Silencing of SOX5 increased GSK3β expression, which indicates a functional link between SOX5 and GSK3β in AD pathogenesis. Silencing of SOX5 also led to an increased expression of two known negative regulators of WNT signaling: Disabled-2 (DAB2) and SOX17. The adaptor molecule DAB2 stabilizes the β-catenin degradation complex. SOX17, a homolog of SOX5, changes β-catenin, SFRP1 and WNT/Frizzled expression. FGF4 is a direct transcriptional target for LEF1 and WNT signaling. It has been shown that FGF infusion or gene transfer restores neurogenesis in the subventricular zone and hippocampal functions in aged mice and mouse models of AD and has therapeutic implications for neurocognitive disorders. WNT11 and WNT2 are components of noncanonical WNT signaling, while WNT9A is the components of canonical WNT signaling (Li, 2017).
Moreover, bone morphogenic proteins (BMPs) as members of the transforming growth factor-β (TGF-β) superfamily are important regulators of neurogenesis and neuronal cell fate determination during development. The hippocampus from either AD patients or APP transgenic mice exhibited significantly increased BMP6 expression accompanied by defects in hippocampal neurogenesis compared to controls. The WNT and the BMP signaling pathways are functionally integrated in many biological processes including stem cell maintenance and cell fate specifications. WNT and BMP ligands are expressed in overlapping or complementary manners. For example, the enhancer of the even-skipped gene (eve) has a BMP response element (Smad1/5/8 and Smad4 binding sites) next to a WNT response element which integrates synergy between WNT and BMP signaling. In mice Sox5 is essential for activation of BMP directed target gene expression in embryos and explants, that it physically interacts with BMP Smad1/5/8, and this interaction is essential for recruitment of Smad1/4 to BMP regulatory elements. Functional studies are warranted to further investigate the regulating effects of SOX5 on genes related to WNT-signaling as well as BMP or TGF-β signaling (Li, 2017).
SOX5 variants have been previously associated with two neurological disorders: ALS and developmental delay or intellectual disability. Two missense mutations in SOX5, Q362P and E367Q, five amino acids apart, were identified in two of the 190 ALS patients and not detected in 190 normal controls. Several mutations including a reciprocal translocation breakpoint within SOX5 and variable deletions in SOX5 that range in size from 72 to 466 kb have been identified in patients with developmental delay or intellectual disability, which are associated with prominent language delay, behavior problems and mild dysmorphic features. In line with the above findings in ALS, this study demonstrates the SOX5 association with LOAD, another neurodegenerative disease attributed to SOX5. Furthermore, four novel functional variants in SOX5 that modify susceptibility to LOAD predicted to have functionality and pathogenicity by computer algorithms annotations. Three of the four novel mutations in the SOX5 gene, two missense mutations SOX5-R36T and H721R and one splice-site variant, segregated with the affection status, suggesting that these three SOX5 variants have an important role in the onset of AD, probably by loss of SOX5 function. In contrast to the deleterious SOX5-R36T and H721R mutations, the AD-linked SOX5-T74M mutation may be acting in a more complex gain-of-function fashion to confer protection from AD risk. Notably, missense mutations in SOX5 identified in LOAD families are clustered in the N and C-termini of SOX5 (R36T, T74M, and H721R); in contrast, the distribution of two missense mutations in ALS patients is in the middle part of SOX5 (Q362P and E367Q). Further studies are warranted to determine the relationship between the specific type and location of SOX5 missense mutations and neuronal development, function and AD neuropathogenesis (Li, 2017).
Structurally SOX5 consists of an HMG domain (542-612 amino-acid) and a pattern of the TonB-dependent receptor protein signature 1 (PS00430) at the N-terminal of the SOX5 protein (1-82 amino-acid). Two of the four identified SOX5 missense mutations (R36T and T74M) are localized in the exon 2 encoding the N-terminal TonB-dependent receptor like region. These data suggest that the TonB-dependent receptor like region in SOX5 may be important in the functional role of SOX5, disruption of which may lead to AD pathogenesis. It is known that the short form of SOX5 differs from the long form in length with a 13 amino acid truncation in N-terminal. Both SOX5-R36T and T74M are located in the N-terminal, which may cause alternative splicing of the SOX5 exon 2 to alter the expression of SOX5 long and/or short isolforms. Notably, SOX5-R36T is associated with an increased AD risk, while SOX5-T74M confers protection from AD risk in three subjects of an AD family who had known increased risk for AD from either homozygous or heterozygous for APOE-ε4. However, the protective mechanism of SOX5-T74M on LOAD, distinct from SOX5-R36T and H721R mutations which increase AD risk, is unclear. One possibility would be if the T74M variant stabilizes SOX5 similar to the PMP22 mutation, T118M, that causes Charcot Marie Tooth, type 1A (CMT1A), by an overall increase in the stability of the mutant PMP22 protein (Li, 2017).
In summary, the findings from this study indicate that SOX5 is a novel LOAD candidate gene with an important role in neuronal function. Furthermore, SOX5 holds significant potential for the presence of additional variants that could confer risk for AD, ALS and intellectual disability (Li, 2017).
The SRY-related HMG-box 5 (SOX5) gene encodes a member of the SOX family of transcription factors. Recently, genome-wide association studies have implicated SOX5 as a candidate gene for susceptibility to four cardiac-related endophenotypes: higher resting heart rate (HR), the electrocardiographic PR interval, atrial fibrillation and left ventricular mass. Human SOX5 has a highly conserved Drosophila ortholog, Sox102F, and have employed transgenic Drosophila models to quantitatively measure cardiac function in adult flies. For this purpose, this study describes a high-speed and ultrahigh-resolution optical coherence tomography imaging system, which enables rapid cross-sectional imaging of the heart tube over various cardiac cycles for the measurement of cardiac structural and dynamical parameters such as HR, dimensions and areas of heart chambers, cardiac wall thickness and wall velocities. The silencing of Sox102F resulted in a significant decrease in HR, heart chamber size and cardiac wall velocities, and a significant increase in cardiac wall thickness that was accompanied by disrupted myofibril structure in adult flies. In addition, the silencing of Sox102F in the wing led to increased L2, L3 and wing marginal veins and increased and disorganized expression of wingless, the central component of the Wnt signaling pathway. Collectively, the silencing of Sox102F resulted in severe cardiac dysfunction and structural defects with disrupted Wnt signaling transduction in flies. This implicates an important functional role for SOX5 in heart and suggests that the alterations in SOX5 levels may contribute to the pathogenesis of multiple cardiac diseases or traits (Li, 2013).
The Drosophila medulla, part of the visual processing center of the brain, contains more than 70 neural types generated by medulla neuroblasts which sequentially express several temporal transcription factors (TTFs), including Homothorax (Hth), eyeless (Ey), Sloppy paired 1 and 2 (Slp), Dichaete (D) and Tailless (Tll). However, it is not clear how such a small number of TTFs could give rise to diverse combinations of neuronal transcription factors that specify a large number of medulla neuron types. This study reports how temporal patterning specifies one neural type, the T1 neuron. The T1 neuron is the only medulla neuron type that expresses the combination of three transcription factors Ocelliless (Oc or Otd), Sox102F and Ets65A. Using CRISPR-Cas9 system, this study shows that each transcription factor is required for the correct morphogenesis of T1 neurons. Interestingly, Oc, Sox102F and Ets65A initiate expression in neurons beginning at different temporal stages and last in a few subsequent temporal stages. Oc expressing neurons are generated in the Ey, Slp and D stages; Sox102F expressing neurons are produced in the Slp and D stages; while Ets65A is expressed in subsets of medulla neurons born in the D and later stages. The TTF Ey, Slp or D is required to initiate the expression of Oc, Sox102F or Ets65A in neurons, respectively. Thus, the neurons expressing all three transcription factors are born in the D stage and become T1 neurons. In neurons where the three transcription factors do not overlap, each of the three transcription factors can act in combination with other neuronal transcription factors to specify different neural fates. This study shows that this way of expression regulation of neuronal transcription factors by temporal patterning can generate more possible combinations of transcription factors in neural progeny to diversify neural fates (Naidu, 2020).
T1 neurons are a class of mysterious neurons that connect the lamina and the medulla part of the optic lobe. They are uni-columnar neurons with one in each of the 800 columns of the medulla. The cell body of the T1 neuron is found in the medulla cortex, and its axon branches in a characteristic 'T' shape on the distal surface of the medulla. One branch projects through the outer optic chiasm back to the lamina and then forms a basket like structure of processes surrounding each lamina cartridge. The other branch arborizes in the M2 layer of the medulla with a dense bush like structure. T1 neuron is post-synaptic to amc (lamina amacrine cells), and the amc/T1 pathway was shown to enhance the lamina neuron L1 motion detection pathway at intermediate contrast. Depolarizing T1 neurons affected the flight steering responses to visual stimuli (Naidu, 2020).
Through screening antibodies and GFP fusion lines, this study found that Ocelliless (oc), Sox102F and Ets65A are expressed in T1 neurons, and the combination of these three TFs can distinguish T1 neurons from all other medulla neurons. Using CRISPR-Cas9 system, bi-allelic somatic mutations of each of the three TF genes were generated in T1 neurons; knock-down of each one affected different aspects of the T1 neuron morphology. Next, how the expression of each TF is controlled by temporal patterning to generate the combination code was examined. Oc expression in neurons starts in the Ey temporal stage, and continues in the Slp and D temporal stages, and Ey is required for the initiation of Oc expression in neurons; while Sox102F expression in neurons starts in the Slp temporal stage, and continues in the D temporal stage, and Slp is required for initiating the expression of Sox102F in neurons; finally, Ets65A is expressed in subsets of medulla neurons born in the D and later temporal stages, and D is required for the expression of Ets65A. Thus, the three TFs that control T1 neuron morphology initiate their expression in neurons beginning at different temporal stages controlled by different TTFs, but each of them spans a few temporal stages, and the neurons expressing all three TFs are born in the D stage and become T1 neurons. In neurons where the three transcription factors do not overlap, each of the three TFs could also act with other neuronal TFs to specify different neural fates. In this way, more combinations of TFs can be generated through temporal patterning (Naidu, 2020).
This study identified a combination of three transcription factors that control T1 neuron morphology, and examined how the expression of these three transcription factors are controlled by temporal patterning of medulla neuroblasts. Oc is turned on in neurons starting in the late Ey stage, and Oc expressing neurons continue to be generated in the Slp and D stages, although the fates will be different, possibly dependent on the co-expression with other neuronal TFs. Sox102F expressing neurons start to be generated in the Slp stage and continue in the D stage. Ets65A expressing neurons are generated in the D and later temporal stages. Thus, the three TFs that control T1 neuron morphology start their expression in neurons born at different temporal stages, and require the corresponding TTF for initiation of their expression, and each neuronal TF is expressed in neurons spanning a few temporal stages. One advantage for such temporal control of neuronal TFs is that more combinations of TFs can be generated to specify different fates. For example, Toy is expressed in the N-on neuronal progeny born from the Slp and D stages, and also in some N-off progeny born from the late Ey stage neuroblasts in some regions of the medulla. Results from this study and others suggest that the subset of Sox102F neurons that do not express Oc, express Toy and Ap instead, and they are specified as Tm5 neurons. In addition, the neurons that express both Toy and Oc in the N-off progeny of some late Ey stage neuroblasts could determine another unknown neural type. Although it remains to be determined whether these TF combinations are indeed required for the corresponding neural fates, these examples do suggest that different combinations of neuronal TFs can be created that might determine different fates (Naidu, 2020).
Mutation of each of the three TFs expressed in T1 caused a certain morphological defect, similar to the morphology TFs that act in combinations to determine motor neuron morphology. For oc and Ets65A mutant neurons, it appeared that they still maintained the T1 fate, but the morphology was abnormal. Some Sox102F mutant neurons resembled medulla intrinsic neurons, but without functional assay, it was not clear whether they were fully transformed to a normal Mi neuron fate, or they still maintained some T1 neuron charateristics but underwent dramatic morphological changes. One question is whether the combination of TFs regulate neuron morphology by simple addition (each TF determines one feature, and the simple addition of these features determines one neural type), or in a synergistic way (three TFs together can determine features not determined by either TF alone). In the case of T1, when Sox102F was removed from T1 neurons, the driver used (T1-LexA) was still expressed in the mutant neurons, but the neurons became more like medulla intrinsic neurons, and some neurons lost the projection back to the lamina. However, Sox102F is not expressed in other neurons that project back to the lamina like lamina wide field neurons (lawf 1/2) which express Hth and Eya. Instead, Sox102F is also expressed in a Transmedulla neural type (Tm5) which do not resemble T1 neurons. Thus, these results favor the synergistic action model of neuronal TFs to control neuron morphology (Naidu, 2020).
The results are consistent with the principle that integration between temporal/spatial patterning of neuroblasts and the Notch-dependent binary neuron fate choice further diversifies neural fates. This study found that T1 neurons are derived from the Notch-off hemilineage of D stage neuroblasts. In addition, although T1 neurons are uni-columnar neurons that are generated throughout the main medulla region, there is a spatial component that regulates Oc expression and neural fate specification. Neurons that co-express Oc and Forkhead are only localized in the Dpp domains. Through analyzing the sequencing data published for all medulla neurons, the neurons expressing both Oc and Fkh should become the Dm12 neuron, a multi-columnar neuron with arborizations spanning several columns. Thus, these results support the conclusion that uni-columnar neurons are generated throughout the medulla main region, while multi-columnar neurons are generated in special spatial domains determined by spatial patterning (Naidu, 2020).
In summary, this study of T1 neuron specification illustrated an example how temporal patterning of neuroblasts sequentially turns on the expression of three TFs in neuronal progeny, and generates different combinational codes to determine neural fates. In the future it will be interesting to examine how TTFs in neuroblasts regulate the expression of neuronal TFs in neurons that often span a few temporal stages. Only a subset of neurons maintain the expression of TTFs, while other neurons do not. Thus the TTFs should determine the expression of neuronal TFs already in neuroblasts. It is possible that the TTF promotes epigenetic modifications in the neuronal TF gene locus, so that the TF will be turned on in its progeny as well as in neurons born in subsequent temporal stages. It is also possible that the expression of the same neuronal TF in two subsequent temporal stages are controlled by two separate enhancers that respond to different TTFs. Addressing these questions will further advance understanding of the link between neuroblast temporal patterning and neural fate specification (Naidu, 2020).
Transcriptional regulation of proteins involved in neuronal polarity is a key process that underlies the ability of neurons to transfer information in the central nervous system. The Collapsin Response Mediator Protein (CRMP) family is best known for its role in neurite outgrowth regulation conducting to neuronal polarity and axonal guidance, including CRMP5 that drives dendrite differentiation. Although CRMP5 is able to control dendritic development, the regulation of its expression remains poorly understood. This study identified a Sox5 consensus binding sequence in the putative promoter sequence upstream of the CRMP5 gene. By luciferase assays Sox5 was shown to increase CRMP5 promoter activity, but not if the putative Sox5 binding site is mutated. Sox5 can physically bind to the CRMP5 promoter DNA in gel mobility shift and chromatin immunoprecipitation assays. Using a combination of real-time RT-PCR and quantitative immunocytochemistry, further evidence is provided for a Sox5-dependent upregulation of CRMP5 transcription and protein expression in N1E115 cells: a commonly used cell line model for neuronal differentiation. Furthermore, increasing Sox5 levels in this neuronal cell line inhibits neurite outgrowth. This inhibition requires CRMP5 because CRMP5 knockdown prevents the Sox5-dependent effect. The physiological relevance of the Sox5-CRMP5 pathway in the regulation of neurite outgrowth was confirmed using mouse primary hippocampal neurons. These findings identify Sox5 as a critical modulator of neurite outgrowth through the selective activation of CRMP5 expression (Naudet, 2018).
Mechanisms generating diverse cell types from multipotent progenitors are fundamental for normal development. Pigment cells are derived from multipotent neural crest cells and their diversity in teleosts provides an excellent model for studying mechanisms controlling fate specification of distinct cell types. Zebrafish have three types of pigment cells (melanocytes, iridophores and xanthophores) while medaka have four (three shared with zebrafish, plus leucophores), raising questions about how conserved mechanisms of fate specification of each pigment cell type are in these fish. Previous work has shown that the Sry-related transcription factor Sox10 is crucial for fate specification of pigment cells in zebrafish, and that Sox5 promotes xanthophores and represses leucophores in a shared xanthophore/leucophore progenitor in medaka. Employing TILLING, TALEN and CRISPR/Cas9 technologies, medaka and zebrafish sox5 and sox10 mutants were generated and comparative analyses of their compound mutant phenotypes was conducted. Specification of all pigment cells, except leucophores, was shown to be dependent on Sox10. Loss of Sox5 in Sox10-defective fish partially rescued the formation of all pigment cells in zebrafish, and melanocytes and iridophores in medaka, suggesting that Sox5 represses Sox10-dependent formation of these pigment cells, similar to their interaction in mammalian melanocyte specification. In contrast, in medaka, loss of Sox10 acts cooperatively with Sox5, enhancing both xanthophore reduction and leucophore increase in sox5 mutants. Misexpression of Sox5 in the xanthophore/leucophore progenitors increased xanthophores and reduced leucophores in medaka. Thus, the mode of Sox5 function in xanthophore specification differs between medaka (promoting) and zebrafish (repressing), which is also the case in adult fish. These findings reveal surprising diversity in even the mode of the interactions between Sox5 and Sox10 governing specification of pigment cell types in medaka and zebrafish, and suggest that this is related to the evolution of a fourth pigment cell type (Nagao, 2018).
In anamniotes, somite compartimentalization in the lateral somitic domain leads simultaneously to myotome and dermomyotome formation. In the myotome, Xenopus Sox5 is co-expressed with Myod1 in the course of myogenic differentiation. The function of Sox5 using a Myod1-induced myogenic transcription assay was studied in pluripotent cells of animal caps. Sox5 was found to enhance myogenic transcription of muscle markers Des, Actc1, Ckm and MyhE3. The use of chimeric transactivating or transrepressive Sox5 proteins indicates that Sox5 acts as a transrepressor and indirectly stimulates myogenic transcription except for the slow muscle-specific genes Myh7L, Myh7S, Myl2 and Tnnc1. This role is shared by Sox6, which is structurally similar to Sox5, both belonging to the SoxD subfamily of transcription factors. Moreover, Sox5 can antagonize the inhibitory function of Meox2 on myogenic differentiation. Meox2 which is a dermomyotome marker, represses myogenic transcription in Myod-induced myogenic transcription assay and in Nodal5-induced mesoderm from animal cap assay. The inhibitory function of Meox2 and the pro-myogenic function of Sox5 were confirmed during Xenopus normal development by the use of translation-blocking oligomorpholinos and dexamethasone inducible chimeric Sox5 and Meox2 proteins. This study has therefore identified a new function for SoxD proteins in muscle cells, which can indirectly enhance myogenic transcription through transrepression, in addition to the previously identified function as a direct repressor of slow muscle-specific genes (Della Gaspera, 2018).
Search PubMed for articles about Drosophila Sox102F or SoxD
Contreras, E. G., Palominos, T., Glavic, A., Brand, A. H., Sierralta, J. and Oliva, C. (2018). The transcription factor SoxD controls neuronal guidance in the Drosophila visual system. Sci Rep 8(1): 13332. PubMed ID: 30190506
Della Gaspera, B., Chesneau, A., Weill, L., Charbonnier, F. and Chanoine, C. (2018). Xenopus SOX5 enhances myogenic transcription indirectly through transrepression. Dev Biol 442(2): 262-275. PubMed ID: 30071218
Li, A., Ahsen, O. O., Liu, J. J., Du, C., McKee, M. L., Yang, Y., Wasco, W., Newton-Cheh, C. H., O'Donnell, C. J., Fujimoto, J. G., Zhou, C. and Tanzi, R. E. (2013). Silencing of the Drosophila ortholog of SOX5 in heart leads to cardiac dysfunction as detected by optical coherence tomography. Hum Mol Genet 22(18): 3798-3806. PubMed ID: 23696452
Li, A., Hooli, B., Mullin, K., Tate, R. E., Bubnys, A., Kirchner, R., Chapman, B., Hofmann, O., Hide, W. and Tanzi, R. E. (2017). Silencing of the Drosophila ortholog of SOX5 leads to abnormal neuronal development and behavioral impairment. Hum Mol Genet 26(8): 1472-1482. PubMed ID: 28186563
Nagao, Y., Takada, H., Miyadai, M., Adachi, T., Seki, R., Kamei, Y., Hara, I., Taniguchi, Y., Naruse, K., Hibi, M., Kelsh, R. N. and Hashimoto, H. (2018). Distinct interactions of Sox5 and Sox10 in fate specification of pigment cells in medaka and zebrafish. PLoS Genet 14(4): e1007260. PubMed ID: 29621239
Naidu, V. G., Zhang, Y., Lowe, S., Ray, A., Zhu, H. and Li, X. (2020). Temporal progression of Drosophila medulla neuroblasts generates the transcription factor combination to control T1 neuron morphogenesis Dev Biol. PubMed ID: 32442418
Naudet, N., Moutal, A., Vu, H. N., Chounlamountri, N., Watrin, C., Cavagna, S., Malleval, C., Benetollo, C., Bardel, C., Dronne, M. A., Honnorat, J., Meissirel, C. and Besancon, R. (2018). Transcriptional regulation of CRMP5 controls neurite outgrowth through Sox5. Cell Mol Life Sci 75(1): 67-79. PubMed ID: 28864883
Pinto-Teixeira, F., Koo, C., Rossi, A. M., Neriec, N., Bertet, C., Li, X., Del-Valle-Rodriguez, A. and Desplan, C. (2018). Development of Concurrent Retinotopic Maps in the Fly Motion Detection Circuit. Cell 173(2): 485-498 e411. PubMed ID: 29576455
date revised: 10 December 2020
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