InteractiveFly: GeneBrief

Myoinhibiting peptide precursor: Biological Overview | References

Gene name - Myoinhibiting peptide precursor

Synonyms - Myoinhibitory peptide

Cytological map position - 74A5-74A5

Function - Secreted peptide

Keywords - ancestral ligand for the Sex peptide receptor, acts directly in the polyamine-detecting olfactory neurons and taste neurons, regulation of polyamine attraction, stabilization of sleep in both males and females

Symbol - Mip

FlyBase ID: FBgn0036713

Genetic map position - chr3L:17,344,947-17,349,666

Classification -

Cellular location - secreted ligand

NCBI link: EntrezGene
Mip orthologs: Biolitmine
Recent literature
Jang, Y. H., Chae, H. S. and Kim, Y. J. (2017). Female-specific myoinhibitory peptide neurons regulate mating receptivity in Drosophila melanogaster. Nat Commun 8(1): 1630. PubMed ID: 29158481
Upon mating, fruit fly females become refractory to further mating for several days. An ejaculate protein called sex peptide (SP) acts on uterine neurons to trigger this behavioural change, but it is still unclear how the SP signal modifies the mating decision. This study describes two groups of female-specific local interneurons that are important for this process-the ventral abdominal lateral (vAL) and ventral abdominal medial (vAM) interneurons. Both vAL and vAM express myoinhibitory peptide (Mip)-GAL4. vAL is positive for Mip neuropeptides and the sex-determining transcriptional factor doublesex. Silencing the Mip neurons in females induces active rejection of male courtship attempts, whereas activation of the Mip neurons makes even mated females receptive to re-mating. vAL and vAM are located in the abdominal ganglion (AG) where they relay the SP signal to other AG neurons that project to the brain. Mip neuropeptides appear to promote mating receptivity both in virgins and mated females, although it is dispensable for normal mating in virgin females.
Reinhard, N., Bertolini, E., Saito, A., Sekiguchi, M., Yoshii, T., Rieger, D. and Helfrich-Forster, C. (2021). The lateral posterior clock neurons of Drosophila melanogaster express three neuropeptides and have multiple connections within the circadian clock network and beyond. J Comp Neurol. PubMed ID: 34961936
Drosophila's lateral posterior neurons (LPNs) belong to a small group of circadian clock neurons that is so far not characterized in detail. Thanks to a new highly specific split-Gal4 line, this study describes LPNs' morphology in fine detail, their synaptic connections, daily bimodal expression of neuropeptides, and a putative role of this cluster in controlling daily activity and sleep patterns is proposed. The three LPNs were found to be heterogeneous. Two of the neurons with similar morphology arborize in the superior medial and lateral protocerebrum and most likely promote sleep. One unique, possibly wakefulness-promoting, neuron with wider arborizations extends from the superior lateral protocerebrum toward the anterior optic tubercle. Both LPN types exhibit manifold connections with the other circadian clock neurons, especially with those that control the flies' morning and evening activity (M- and E-neurons, respectively). In addition, they form synaptic connections with neurons of the mushroom bodies, the fan-shaped body, and with many additional still unidentified neurons. Both LPN types rhythmically express three neuropeptides, Allostatin A, Allostatin C, and Diuretic Hormone 31 with maxima in the morning and the evening. The three LPN neuropeptides may, furthermore, signal to the insect hormonal center in the pars intercerebralis and contribute to rhythmic modulation of metabolism, feeding, and reproduction. These findings are discussed in the light of anatomical details gained by the recently published hemibrain of a single female fly on the electron microscopic level and of previous functional studies concerning the LPN.
Thoma, V., Sakai, S., Nagata, K., Ishii, Y., Maruyama, S., Abe, A., Kondo, S., Kawata, M., Hamada, S., Deguchi, R. and Tanimoto, H. (2023). On the origin of appetite: GLWamide in jellyfish represents an ancestral satiety neuropeptide. Proc Natl Acad Sci U S A 120(15): e2221493120.. PubMed ID: 37011192
Food intake is regulated by internal state. This function is mediated by hormones and neuropeptides, which are best characterized in popular model species. However, the evolutionary origins of such feeding-regulating neuropeptides are poorly understood. The jellyfish Cladonema was used to address this question. Combined transcriptomic, behavioral, and anatomical approaches identified GLWamide as a feeding-suppressing peptide that selectively inhibits tentacle contraction in this jellyfish. In the fruit fly Drosophila, myoinhibitory peptide (MIP) is a related satiety peptide. Surprisingly, it was found that GLWamide and MIP were fully interchangeable in these evolutionarily distant species for feeding suppression. These results suggest that the satiety signaling systems of diverse animals share an ancient origin.

A female's reproductive state influences her perception of odors and tastes along with her changed behavioral state and physiological needs. The mechanism that modulates chemosensory processing, however, remains largely elusive. Using Drosophila, this study has identified a behavioral, neuronal, and genetic mechanism that adapts the senses of smell and taste, the major modalities for food quality perception, to the physiological needs of a gravid female. Pungent smelling polyamines, such as putrescine and spermidine, are essential for cell proliferation, reproduction, and embryonic development in all animals. A polyamine-rich diet increases reproductive success in many species, including flies. Using a combination of behavioral analysis and in vivo physiology, this study shows that polyamine attraction is modulated in gravid females through a G-protein coupled receptor, the sex peptide receptor (SPR), and its neuropeptide ligands, MIPs (myoinhibitory peptides), which act directly in the polyamine-detecting olfactory and taste neurons. This modulation is triggered by an increase of SPR expression in chemosensory neurons, which is sufficient to convert virgin to mated female olfactory choice behavior. Together, these data show that neuropeptide-mediated modulation of peripheral chemosensory neurons increases a gravid female's preference for important nutrients, thereby ensuring optimal conditions for her growing progeny (Hussain, 2016b).

The behavior of females in most animal species changes significantly as a consequence of mating. Those changes are interpreted from an evolutionary standpoint as the female's preparation to maximize the fitness of her offspring. In general, they entail a qualitative and quantitative change in her diet, as well as the search for an optimal site where her progeny will develop. In humans, the eating behavior and perception of tastes and odors of a pregnant woman are modulated in concert with altered physiology and the specific needs of the embryo. While several neuromodulatory molecules such as noradrenaline are found in the vertebrate olfactory and gustatory systems, little is known about how reproductive state and pregnancy shape a female's odor and taste preferences. Very recent work in the mouse showed that olfactory sensory neurons (OSNs) are modulated during the estrus cycle (Dey, 2015). Progesterone receptor expressed in OSNs decreases the sensitivity of pheromone-detecting OSNs and thereby reduces the non-sexually receptive female's interest in male pheromones. The mechanisms of how mating, pregnancy, and lactation shape the response of the female olfactory and gustatory systems remain poorly understood (Hussain, 2016b).

The neuronal underpinnings of mating and its consequences on female behaviors have arguably been best characterized in Drosophila. Shortly after copulation, female flies engage in a series of post-mating behaviors contrasting with those of virgins: their sexual receptivity decreases, and they feed to accumulate essential resources needed for the production of eggs; finally, they lay their eggs. This suite of behaviors results from a post-mating trigger located in the female's reproductive tract (Kubli, 2003). Sensory neurons extending their dendrites directly into the oviduct are activated by a component of the male's ejaculate, the sex peptide (SP) (Hasemeyer, 2009; Yang, 2009). Sex peptide receptor (SPR) expressed by these sensory neurons triggers the post-mating switch. Mated females mutant for SPR produce and lay fewer eggs while maintaining a high sexual receptivity. In addition to SP, male ejaculate contains more than 200 proteins, which are transferred along with SP into the female. These have been implicated in conformational changes of the uterus, induction of ovulation, and sperm storage (Hussain, 2016b).

Additional SPR ligands have been identified that are not required for the canonical post-mating switch, opening the possibility that this receptor is involved in the neuromodulation of other processes (Kim, 2010; Isaac, 2014; Hull, 2014; Conzelmann, 2013). These alternative ligands, the myoinhibitory peptides (MIPs)/allatostatin-Bs, unlike SP, have been found outside of drosophilids, in many other insect species such as the silkmoth (Bombyx mori), several mosquito species, and the red flour beetle (Tribolium castaneum). They are expressed in the brain of flies and mosquitoes, including in the centers of olfactory and gustatory sensory neuron projections, the antennal lobe (AL), and the subesophageal zone (SEZ), respectively (Kim, 2010; Siju, 2014; Carlsson, 2010). Although these high-affinity SPR ligands have recently been implicated in the control of sleep in Drosophila males and females, nothing thus far suggests a function in reproductive behaviors (Hussain, 2016b).

To identify optimal food and oviposition sites, female flies rely strongly on their sense of smell and taste. Drosophila females prefer to oviposit in decaying fruit and use byproducts of fermentation such as ethanol and acetic acid to choose oviposition sites. Their receptivity to these byproducts is enhanced by their internal state. It was shown, for instance, that the presence of an egg about to be laid results in increased attraction to acetic acid. Yet the mechanisms linking reproductive state to the modulation of chemosensory processing remain unknown (Hussain, 2016b).

This study has examined the causative mechanisms that integrate reproductive state into preference behavior and chemosensory processing. Focus was placed on the perception of another class of byproducts of fermenting fruits, polyamines. Polyamines such as putrescine, spermine, and spermidine are important nutrients that are associated with reproductive success across animal species (Lefèvre, 2011). A diet high in polyamines indeed increases the number of offspring of a fly couple, and female flies prefer to lay their eggs on polyamine-rich food (Hussain, 2016a). Importantly, previous studies have characterized the chemosensory mechanisms flies use to find and evaluate polyamine-rich food sources and oviposition sites. In brief, volatile polyamines are detected by OSNs on the fly's antenna, co-expressing two ionotropic receptors (IRs), IR41a and IR76b (Hussain, 2016a; Silbering, 2011). Interestingly, the taste of polyamines is also detected by IR76b in labellar gustatory receptor neurons (GRNs) (Hussain 2016a; Hussain, 2016b).

This beneficial role of polyamines has a well-characterized biological basis: polyamines are essential for basic cellular processes such as cell growth and proliferation, and are of specific importance during reproduction. They enhance the quality of sperm and egg and are critical during embryogenesis and postnatal development. While the organism can generate polyamines, a significant part is taken in with the diet. Moreover, endogenous synthesis of polyamines declines with ageing and can be compensated for through a polyamine-rich diet. Therefore, these compounds represent a sensory cue as well as an essential component of the diet of a gravid female fly (Hussain, 2016b and references therein).

This study shows that the olfactory and gustatory perception of polyamines is modulated by the female's reproductive state and guides her choice behavior accordingly. This sensory and behavioral modulation depends on SPR and its conserved ligands, the MIPs that act directly on the chemosensory neurons themselves. Together, these results suggest that mating-state-dependent neuropeptidergic modulation of chemosensory neurons matches the female fly's decision-making to her physiological needs (Hussain, 2016b).

Mechanistically, this study shows that virgin females, or mated females lacking the G-protein coupled receptor SPR, display reduced preference for polyamine-rich food and oviposition sites. Using targeted gene knockdown, mutant rescue, overexpression, and in vivo calcium imaging, a new role was uncovered for SPR and its conserved ligands, MIPs, in directly regulating the sensitivity of chemosensory neurons and modulating taste and odor preferences according to reproductive state. Together with recent work in the mouse (Dey, 2015), these results emphasize that chemosensory neurons are potent targets for tuning choice behavior to reproductive state (Hussain, 2016b).

Reproductive behaviors such as male courtship and female egg-laying strongly depend on the mating state. While previous work has suggested that mating modulates odor- or taste-driven choice behavior of Drosophila females, how mating changes the processing of odors and tastes remained elusive. This study shows that a female-specific neuropeptidergic mechanism acts in peripheral chemosensory neurons to enhance female preference for essential nutrients. The data suggests that this modulation is autocrine and involves the GPCR SPR and its conserved MIP ligands. Notably, MIPs are expressed in chemosensory cells in the apical organs of a distant organism, the annelid (Platynereis) larvae, in which they trigger settlement behavior via an SPR-dependent signaling cascade (Conzelmann, 2013). Importantly, as SP and not MIP induces the SPR-dependent canonical post-mating switch, the current findings report the first gender and mating-state-dependent role of these peptides (Oh, 2014). Whether this regulation is also responsible for previously reported changes in preference behavior upon mating remains to be seen, but it is anticipated that this type of regulation is not only specific to polyamines. On the other hand, mating-dependent changes for salt preference-salt preference is also dependent on IR76b receptor but in another GRN type-might undergo a different type of regulation, as RNAi-mediated knockdown of SPR in salt receptor neurons had no effect on salt feeding (Walker, 2015). Instead, the change in salt preference is mediated by the canonical SP/SPR pathway and primarily reflects the fact that mating has taken place. The mechanism of how salt detection and/or processing are modulated is not known. In contrast to salt preference and polyamine preference, acetic acid preference is strongly modulated by egg-laying activity and not just mating (Gou, 2014). The extent to which changes in salt or acetic acid preference are similar to the modulation of behavior to polyamine that this study has described can currently not be tested, because the olfactory neurons that mediate acetic acid preference have not been determined (Hussain, 2016b).

While SPR regulates the neuronal output of both olfactory and gustatory neurons, the behavioral and physiological data surprisingly revealed that it does so through two opposite neuronal mechanisms. SPR signaling increases the presynaptic response of GRNs and decreases it in OSNs. Behaviorally, these two types of modulation produce the same effect: they enhance the female's attraction to polyamine and tune it to levels typical for decaying or fermenting fruit. How these two effects are regulated by the same receptor and ligand pair remains open. GPCRs can recruit and activate different G-proteins. SPR was previously shown to recruit the inhibitory Gαi/o-type, thereby down-regulating cAMP levels in the cell. In the female reproductive tract, SP inhibits SPR-expressing internal sensory neurons and thereby promotes several post-mating behaviors. This type of inhibitory G-protein signaling could also explain the data in the olfactory system. Here, mating decreases the presynaptic activity of polyamine-detecting OSNs, and conversely, RNAi knockdown of SPR increases their responses strongly. This decrease in neuronal output also shifts the behavioral preference from low to high polyamine levels. While the relationship between behavior and GRN activity is much more straightforward in the gustatory system (increased neuronal response, increased preference behavior), it implies that another G-protein might be activated downstream of SPR. G-protein Gαi/s increases cAMP levels and Gαq enhances phospholipase C (PLC) and calcium signaling. In addition, Gβγ subunits regulate ion channels and other signaling effectors, including PLC. Future work will address the exact mechanisms of this bi-directional modulation through SPR signaling. Nonetheless, it is interesting to speculate that different cells, including sensory neurons, could be modulated differentially by the same molecules depending on cell-specific states and the availability of signaling partners (Hussain, 2016b).

While the data provides a neuronal and molecular mechanism of how chemosensory processing itself is affected by mating, it remains unclear how mating regulates MIP/SPR signaling in chemosensory neurons. The data indicates that SPR levels strongly increase in chemosensory organs upon mating. In addition, MIP levels appear to be mildly increased by mating. This suggests that mating regulates primarily the expression of the GPCR resembling the modulation of sNPFR expression during hunger states. On the other hand, MIP overexpression also induced mated-like preference behavior in virgin flies, suggesting a somewhat more complex situation. For instance, it is possible that overexpression of MIP induces the expression of SPR. Alternatively, active MIP levels might also be regulated at the level of secretion or posttranslational processing, and overexpression might override this form of regulation. In the case of hunger, sNPFR levels are increased through a reduction of insulin signaling. SP could be viewed as the possible equivalent of insulin for mating state. Females mated to SP mutant males, however, do not show a significant change in olfactory perception of polyamines. It is yet important to note that male sperm contains roughly 200 different proteins, some of which might be involved in mediating the change in MIPs/SPR signaling upon mating. In the mosquito, which does not possess SP, the steroid hormone 20E serves as the post-mating switch. Interestingly, mating or treatment with 20E induces in particular the expression of the enzymes required for the synthesis of polyamines in the female spermatheca, a tissue involved in sperm storage and egg-laying. Whether such a mechanism also exists in Drosophila is not known (Hussain, 2016b).

In addition to mating and signals transferred by mating, it is possible that egg-laying activity contributes to the regulation of MIPs/SPR signaling in chemosensory neurons through a mechanism that involves previously identified mechanosensory neurons of the female's reproductive tract; such neurons may sense the presence of an egg to be laid. Indeed, females that cease to lay eggs return to polyamine preferences as found before mating. On the other hand, SP mutant male-mated females and ovoD1 sterile females still show enhanced attraction to polyamine odor, although they lay very few or no eggs. Conversely, knockdown of SPR in IR41a neurons reduced polyamine odor attraction but had a marginal effect on the number of eggs laid. Nevertheless, somewhat reduced numbers of eggs laid were observed upon inactivation of IR76b neurons. At this point, possible reasons can only be speculated. Although IR76b receptor is not expressed in ppk-positive internal SPR neurons, no expression of IR76b-Gal4 is found in neurons innervating the rectum and possibly gut. Hence, there might be an IR76b-mediated interplay between metabolism and nutrient uptake that influences egg-laying. However, females mated to SP-mutant males do not display an increase in feeding, indicating that preference for polyamines does not depend on the metabolic cost of egg-laying. This conclusion is strengthened by the data obtained with mated ovoD1 sterile females, who show similar attraction to polyamines as compared to mated controls. Due to very few or no eggs laid by SP mutant male-mated females and ovoD1 females, respectively, it is not possible to fully exclude a contribution of egg-laying activity to taste-dependent oviposition choice behavior (Hussain, 2016b).

A further argument against an important role of egg-laying activity in the current paradigm comes from the observation that the sensory modulation of OSNs and GRNs occurs rapidly after mating and is maintained only for a few hours. Similarly, SPR expression increases within the same time window shortly after mating. Egg-laying, however, continues for several days after this. In addition, overexpression of SPR was sufficient to switch virgin OSN calcium responses and female behavioral preferences to that of mated females without increasing the number of eggs laid. All in all, these data are more consistent with the hypothesis that mating and not egg-laying activity per se is the primary inducer of sensory modulation leading to the behavioral changes of females (Hussain, 2016b).

It remains that the exact signal triggered by mating that regulates odor and taste preference for polyamines, through the mechanism identified in this study, needs to still be determined. Furthermore, the role of metabolic need and polyamine metabolism is not completely clear. This is similar to the situation found for increased salt preference after mating. While more salt is beneficial for egg-laying, sterile females still increase their preference for salt upon mating. Regardless, in the case of polyamines, it is tempting to speculate that exogenous (by feeding) and endogenous (by enzymatic activity or expression) polyamine sources are regulated by reproductive state and together contribute to reach optimal levels for reproduction in the organism. (Hussain, 2016b).

The results bear some similarities to recent work on the modulation of OSN sensitivity in hunger states (Root, 2011). sNPF/sNPFR signaling modulates the activity of OSNs in the hungry fly. MIPs/SPR might play a very similar role in the mated female. Overexpression of sNPFR in OSNs of fed flies was sufficient to trigger enhanced food search behavior (Root, 2011). Likewise, an increase in SPR signaling in taste or smell neurons converts virgin to mated female preference behavior. Therefore, different internal states appear to recruit similar mechanisms to tune fly behavior to internal state. Furthermore, such modulation of first order sensory neurons appears not only be conserved within a species, but also for regulation of reproductive state-dependent behavior across species. For instance, a recent study in female mice showed that progesterone-receptor signaling in OSNs modulates sensitivity and behavior to male pheromones according to the estrus cycle. Also in this case, sensory modulation accounts in full for the switch in preference behavior. What is the biological significance of integrating internal state at the level of the sensory neuron? First, silencing of neurons in a state-dependent manner shields the brain from processing unnecessary information. As sensory information may not work as an on/off switch, it is possible that an early shift in neural pathway activation might reduce costly inhibitory activity to counteract activation once the sensory signal has been propagated. Second, another interesting possibility is that peripheral modulation might help to translate transient changes in internal state into longer-lasting behavioral changes that manifest in higher brain centers. This might be especially important in the case of female reproductive behaviors such as mate choice or caring for pups or babies. In contrast to hunger, which increases with time of starvation, the effect of mating decays slowly over time as the sperm stored in the female's spermatheca is used up. This study has shown that the effect of mating on chemosensory neurons mediated by MIPs/SPR signaling is strong within the first 6 h after mating and remains a trend at 1 wk post-mating. However, it triggers a long-lasting behavioral switch, which is observed for over a week. Therefore, this transient modulation and altered sensitivity to polyamines could be encoded more permanently in the brain when the animal encounters the stimulus, for instance, in the context of an excellent place to lay her eggs. Because polyamine preference continues to be high for as long as stored sperm can fertilize the eggs, it is speculated that this change in preference might be maintained by a memory mechanism in higher centers of chemosensory processing. Thus, short-term sensory enhancement not only increases perceived stimulus intensity, it may also help to associate a key sensation to a given reward or punishment. These chemosensory associations are of critical importance in parent-infant bonding in mammals, including humans, which form instantly after birth and last for months, years, or a lifetime (Hussain, 2016b).

Control of RUNX-induced repression of Notch signaling by MLF and its partner DnaJ-1 during Drosophila hematopoiesis
In Drosophila, Myeloid Leukemia Factor (MLF) has been shown to control blood cell development by stabilizing the RUNX transcription factor Lozenge (Lz). This study further characterized MLF's mode of action in Drosophila blood cells using proteomic, transcriptomic and genetic approaches. The results show that MLF and the Hsp40 co-chaperone family member DnaJ-1 interact through conserved domains and demonstrate that both proteins bind and stabilize Lz in cell culture, suggesting that MLF and DnaJ-1 form a chaperone complex that directly regulates Lz activity. Importantly, dnaj-1 loss causes an increase in Lz+ blood cell number and size similarly as in mlf mutant larvae. Moreover dnaj-1 was found to genetically interact with mlf to control Lz level and Lz+ blood cell development in vivo. In addition, mlf and dnaj-1 loss was shown to alter Lz+ cell differentiation, and the increase in Lz+ blood cell number and size observed in these mutants is caused by an overactivation of the Notch signaling pathway. Finally, high levels of Lz were shown to be required to repress Notch transcription and signaling. These data indicate that the MLF/DnaJ-1-dependent increase in Lz level allows the repression of Notch expression and signaling to prevent aberrant blood cell development. Thus these findings establish a functional link between MLF and the co-chaperone DnaJ-1 to control RUNX transcription factor activity and Notch signaling during blood cell development in vivo (Miller, 2017).

Proper blood cell development requires the finely tuned regulation of transcription factors and signaling pathways activity. Consequently mutations affecting key regulators of hematopoiesis such as members of the RUNX transcription factor family or components of the Notch signaling pathway are associated with several blood cell disorders including leukemia. Also, leukemic cells often present recurrent chromosomal rearrangements that participate in malignant transformation by altering the function of these factors. The functional characterization of these genes is thus of importance not only to uncover the molecular basis of leukemogenesis but also to decipher the regulatory mechanisms controlling normal blood cell development. Myeloid Leukemia Factor 1 (MLF1) was identified as a target of the t(3;5)(q25.1;q34) translocation associated with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) more than 20 years ago. Further findings suggested that MLF1 could act as an oncogene or a tumor suppressor depending on the cell context and it was shown that MLF1 overexpression either impairs cell cycle exit and differentiation, promotes apoptosis, or inhibits proliferation in different cultured cell lines. Yet, its function and mechanism of action remain largely unknown (Miller, 2017).

MLF1 is the founding member of a small evolutionarily conserved family of nucleo-cytoplasmic proteins present in all metazoans but lacking recognizable domains that could help define their biochemical activity . Whereas vertebrates have two closely related MLF paralogs, Drosophila has a single mlf gene encoding a protein that displays around 50% identity with human MLF in the central conserved domain. In the fly, MLF was identified as a partner of the transcription factor DREF (DNA replication-related element-binding factor), for which it acts a co-activator to stimulate the JNK pathway and cell death in the wing disc. MLF has been shown to bind chromatin, as does its mouse homolog, and it can either activate or repress gene expression by a still unknown mechanism. MLF also interacts with Suppressor of Fused, a negative regulator of the Hedgehog signaling pathway, and, like its mammalian counterpart, with Csn3, a component of the COP9 signalosome, but the functional consequences of these interactions remain elusive. Interestingly the overexpression of Drosophila MLF or that of its mammalian counterparts can suppress polyglutamine-induced cytotoxicity in fly and in cellular models of neurodegenerative diseases. Moreover phenotypic defects associated with MLF loss in Drosophila can be rescued by human MLF1. Thus MLF function seems conserved during evolution and Drosophila appears to be a genuine model organism to characterize MLF proteins (Miller, 2017).

Along this line, the role of MLF during Drosophila hematopoiesis has been studied. Indeed, a number of proteins regulating blood cell development in human, such as RUNX and Notch, also control Drosophila blood cell development. In Drosophila, the RUNX factor Lozenge (Lz) is specifically expressed in crystal cells and it is absolutely required for the development of this blood cell lineage. Crystal cells account for ±4% of the circulating larval blood cells; they are implicated in melanization, a defense response related to clotting, and they release their enzymatic content in the hemolymph by bursting. The Notch pathway also controls the development of this lineage: it is required for the induction of Lz expression and it contributes to Lz+ cell differentiation as well as to their survival by preventing their rupture. Interestingly, the previous analysis revealed a functional and conserved link between MLF and RUNX factors. In particular, MLF was shown to control Lz activity and prevent its degradation in cell culture, and the regulation of Lz level by MLF is critical to control crystal cell number in vivo. Intriguingly, although Lz is required for crystal cell development, mlf mutation causes a decrease in Lz expression but an increase in crystal cell number. In human, the deregulation of RUNX protein level is associated with several pathologies. For instance haploinsufficient mutations in RUNX1 are linked to MDS/AML in the case of somatic mutations, and to familial platelet disorders associated with myeloid malignancy for germline mutations. In the opposite, RUNX1 overexpression can promote lymphoid leukemia. Understanding how the level of RUNX protein is regulated and how this affects specific developmental processes is thus of particular importance (Miller, 2017).

To better characterize the function and mode of action of MLF in Drosophila blood cells, this study used proteomic, transcriptomic and genetic approaches. In line with recent findings, MLF was found to bind DnaJ-1, a HSP40 co-chaperone, as well as the HSP70 chaperone Hsc70-4, and that both of these proteins are required to stabilize Lz. It was further shown that MLF and DnaJ-1 interact together but also with Lz via conserved domains and that they regulate Lz-induced transactivation in a Hsc70-dependent manner in cell culture. In addition, using a null allele of dnaj-1, it was shown to control Lz+ blood cell number and differentiation as well as Lz activity in vivo in conjunction with mlf. Notably, w mlf or dnaj-1 loss leads to an increase in Lz+ cell number and size due to the over-activation of the Notch signaling pathway. Interestingly, these results indicate that high levels of Lz are required to repress Notch expression and signaling. A model is proposed whereby MLF and DnaJ-1 control Lz+ blood cell growth and number by promoting Lz accumulation, which ultimately turndowns Notch signaling. These findings thus establish a functional link between the MLF/Dna-J1 chaperone complex and the regulation of a RUNX-Notch axis required for blood cell homeostasis in vivo (Miller, 2017).

Members of the RUNX and MLF families have been implicated in the control of blood cell development in mammals and Drosophila and deregulation of their expression is associated with human hemopathies including leukemia. The current results establish the first link between the MLF/DnaJ-1 complex and the regulation of a RUNX transcription factor in vivo. In addition, these data show that the stabilization of Lz by the MLF/DnaJ-1 complex is critical to control Notch expression and signaling and thereby blood cell growth and survival. These findings pinpoint the specific function of the Hsp40 chaperone DnaJ-1 in hematopoiesis, reveal a potentially conserved mechanism of regulation of RUNX activity and highlight a new layer of control of Notch signaling at the transcriptional level (Miller, 2017).

MLF binds DnaJ-1 and Hsc70-4, and these two proteins, like MLF, are required for Lz stable expression in Kc167 cells. In addition, these data show that MLF and DnaJ-1 bind to each other via evolutionarily conserved domains and also interact with Lz, suggesting that Lz is a direct target of a chaperone complex formed by MLF, DnaJ-1 and Hsc70-4. Of note, a systematic characterization of Hsp70 chaperone complexes in human cells identified MLF1 and MLF2 as potential partners of DnaJ-1 homologs, DNAJB1, B4 and B6, a finding corroborated by Dyer (2017). Therefore, the MLF/DnaJ-1/Hsc70 complex could play a conserved role in mammals, notably in the regulation of the stability of RUNX transcription factors. How MLF acts within this chaperone complex remains to be determined. In vivo, this study demonstrated that dnaj-1 mutations lead to defects in crystal cell development strikingly similar to those observed in mlf mutant larvae, and these two genes were shown to act together to control Lz+ cells development by impinging on Lz activity. The data suggest that in the absence of DnaJ-1, high levels of MLF lead to the accumulation of defective Lz protein whereas lower levels of MLF allow its degradation. Thus it is proposed that MLF stabilizes Lz and, together with DnaJ-1, promotes its proper folding/conformation. In humans, DnaJB4 stabilizes wild-type E-cadherin but induces the degradation of mutant E-cadherin variants associated with hereditary diffuse gastric cancer. Thus the fate of DnaJ client proteins is controlled at different levels and MLF might be an important regulator in this process (Miller, 2017).

This work presents the first null mutant for a gene of the DnaJB family in metazoans and the results demonstrate that a DnaJ protein is required in vivo to control hematopoiesis. There are 16 DnaJB and in total 49 DnaJ encoding genes in mammals and the expansion of this family has likely played an important role in the diversification of their functions. DnaJB9 overexpression was found to increase hematopoietic stem cell repopulation capacity and Hsp70 inhibitors have anti-leukemic activity, but the participation of other DnaJ proteins in hematopoiesis or leukemia has not been explored. Actually DnaJ's molecular mechanism of action has been fairly well studied but there are only limited insights as to their role in vivo. Interestingly though, both DnaJ-1 and MLF suppress polyglutamine protein aggregation and cytotoxicity in Drosophila models of neurodegenerative diseases, and this function is conserved in mammals. It is tempting to speculate that MLF and DnaJB proteins act together in this process as well as in leukemogenesis. Thus a better characterization of their mechanism of action may help develop new therapeutic approaches for these diseases (Miller, 2017).

As shown in this study, mlf or dnaj-1 mutant larvae harbor more crystal cells than wild-type larvae. This rise in Lz+ cell number is not due to an increased induction of crystal cell fate as we could rescue this defect by re-expressing DnaJ-1 or Lz with the lz-GAL4 driver, which turns on after crystal cell induction, and it was also observed in lz hypomorph mutants, which again suggests a post-lz / cell fate choice process. Moreover mlf or dnaj-1 mutant larvae display a higher fraction of the largest lz>GFP+ cell population, which could correspond to the more mature crystal cells. It is thus tempting to speculate that mlf or dnaj-1 loss promotes the survival of fully differentiated crystal cells. RNAseq data demonstrate that mlf is critical for expression of crystal cell associated genes, but both up-regulation and down-regulation of crystal cell differentiation markers were observed in mlf or dnaj-1 mutant Lz+ cells. Also these changes did not appear to correlate with crystal cell maturation status since alterations were found in gene expression in the mutants both in small and large Lz+ cells. In addition the transcriptome did not reveal a particular bias toward decreased expression for 'plasmatocyte' markers in Lz+ cells from mlf- mutant larvae. Thus, it appears that MLF and DnaJ-1 loss leads to the accumulation of mis-differentiated crystal cells (Miller, 2017).

The data support a model whereby MLF and DnaJ-1 act together to promote Lz accumulation, which in turn represses Notch transcription and signaling pathway to control crystal cell size and number. Indeed, an abnormal maintenance of Notch expression was observed in the larger Lz+ cells as well as an over-activation of the Notch pathway in the crystal cell lineage of mlf and dnaj-1 mutants or when Lz activity was interfered with. Moreover the data as well as previously published experiments show that Notch activation promotes crystal cell growth and survival. Importantly too the increase in Lz+ cell number and size observed in mlf or dnaJ-1 mutant is suppressed when Notch dosage is decreased. Yet, some of the mis-differentiation phenotypes in the mlf or dnaj-1 mutants might be independent of Notch since changes in crystal cell markers expression seem to appear before alterations in Notch are apparent. At the molecular level, the results suggest that Lz directly represses Notch transcription as this study identified a Lz-responsive Notch cis-regulatory element that contains conserved RUNX binding sites. The activation of the Notch pathway in circulating Lz+ cells is ligand-independent and mediated through stabilization of the Notch receptor in endocytic vesicles. Hence a tight control of Notch expression is of particular importance to keep in check the Notch pathway and prevent the abnormal development of the Lz+ blood cell lineage. Notably, Notch transcription was shown to be directly activated by Notch signaling. Such an auto-activation loop might rapidly go awry in a context in which Notch pathway activation is independent of ligand binding. By promoting the accumulation of Lz during crystal cell maturation, MLF and DnaJ-1 thus provide an effective cell-autonomous mechanism to inhibit Notch signaling. Further experiments will now be required to establish how Lz represses Notch transcription. RUNX factors can act as transcriptional repressors by recruiting co-repressor such as members of the Groucho family. Whether MLF and DnaJ-1 directly contribute to Lz-induced-repression in addition to regulating its stability is an open question. MLF and DnaJ-1 were recently found to bind and regulate a common set of genes in cell culture. They may thus provide a favorable chromatin environment for Lz binding or be recruited with Lz and/or favor a conformation change in Lz that allows its interaction with co-repressors. The scarcity of lz>GFP+ cells precludes a biochemical characterization of Lz, MLF and DnaJ-1 mode of action notably at the chromatin level but further genetic studies should help decipher their mode of action. While the post-translational control of Notch has been extensively studied, its transcriptional regulation seems largely overlooked. The current findings indicate that this is nonetheless an alternative entry point to control the activity of this pathway. Given the importance of RUNX transcription factor and Notch signaling in hematopoiesis and blood cell malignancies, it will be of particular interest to further study whether RUNX factors can regulate Notch expression and signaling during these processes in mammals (Miller, 2017).

Myeloid Leukemia Factor acts in a chaperone complex to regulate transcription factor stability and gene expression

Mutations that affect myelodysplasia/myeloid leukemia factor (MLF) proteins are associated with leukemia and several other cancers. However, with no strong homology to other proteins of known function, the role of MLF proteins in the cell has remained elusive. This study describes a proteomics approach that identifies MLF as a member of a nuclear chaperone complex containing a DnaJ protein, BCL2-associated anthanogene 2, and Hsc70. This complex associates with chromatin and regulates the expression of target genes. The MLF complex is bound to sites of nucleosome depletion and sites containing active chromatin marks (e.g., H3K4me3 and H3K4me1). Hence, MLF binding is enriched at promoters and enhancers. Additionally, the MLF-chaperone complex functions to regulate transcription factor stability, including the RUNX transcription factor involved in hematopoiesis. Although Hsc70 and other co-chaperones have been shown to play a role in nuclear translocation of a variety of proteins including transcription factors, these findings suggest that MLF and the associated co-chaperones play a direct role in modulating gene transcription (Dyer, 2017).

A homeostatic sleep-stabilizing pathway in Drosophila composed of the Sex Peptide receptor and its ligand, the myoinhibitory peptide

Sleep, a reversible quiescent state found in both invertebrate and vertebrate animals, disconnects animals from their environment and is highly regulated for coordination with wakeful activities, such as reproduction. The fruit fly, Drosophila melanogaster, has proven to be a valuable model for studying the regulation of sleep by circadian clock and homeostatic mechanisms. This study demonstrates that the Sex peptide receptor (SPR) of Drosophila, known for its role in female reproduction, is also important in stabilizing sleep in both males and females. Mutants lacking either the SPR or its central ligand, myoinhibitory peptide (MIP), fall asleep normally, but have difficulty in maintaining a sleep-like state. This analyses have mapped the SPR sleep function to pigment dispersing factor (pdf) neurons, an arousal center in the insect brain. MIP downregulates intracellular cAMP levels in pdf neurons through the SPR. MIP is released centrally before and during night-time sleep, when the sleep drive is elevated. Sleep deprivation during the night facilitates MIP secretion from specific brain neurons innervating pdf neurons. Moreover, flies lacking either SPR or MIP cannot recover sleep after the night-time sleep deprivation. These results delineate a central neuropeptide circuit that stabilizes the sleep state by feeding a slow-acting inhibitory input into the arousal system and plays an important role in sleep homeostasis (Oh, 2014).

This study reports the discovery of a peptidergic modulatory pathway particularly important in stabilizing sleep and maintaining sleep homeostasis in Drosophila. The key molecules in this novel sleep-regulating pathway are SPR and its peptide ligand MIP. SPR was first identified as a receptor that triggers post-mating behavioral responses (PMR) by mediating actions of the seminal protein SP in females. Although previous biochemical studies demonstrated that SPR could interact with MIP as well as SP, there was no evidence that the interaction between MIP and SPR is biologically relevant in Drosophila (Kim, 2010). By combining genetic analyses and optical activity imaging, this study has provided several independent lines of evidence demonstrating that MIP consolidates sleep state and maintain sleep homeostasis by acting through SPR expressed in arousal-promoting pdf neurons (Oh, 2014).

First, flies lacking either SPR or MIP have a highly similar sleep phenotype. Second, sleep phenotypes of MIP or SPR mutant are manifested regardless of sex, consistent with previous accounts that unlike SP, MIP and SPR expression in the brain show little sexual difference. Third, ex vivo optical activity imaging revealed that exogenous application of MIP downregulates cAMP levels in SPR-expressing pdf neurons, but not in SPR-deficient mutant neurons. Fourth, the sleep phenotypes of SPR-deficient mutants are rescued by restoring SPR expression with insect SPRs that are highly sensitive to MIP, but less sensitive to SP. Hence, SPR interacts with two evolutionarily unrelated sets of ligands, each of which controls completely different behaviors: SP for reproductive behaviors and MIP for sleep behavior. For sleep behaviors, all phenotypes observed in the SPR deficient mutant were also observed in MIP-RNAi. Thus, there is no reason to assume additional ligand(s) for SPR besides SP and MIP at this moment. Nevertheless, the finding that a peptide GPCR can mediate actions of multiple, unrelated groups of ligands should be taken into consideration in searching for peptides and/or other types of ligands for GPCRs (Oh, 2014).

Genetic analyses demonstrated that the expression of SPR in three Gal4 neural populations (cry-Gal4, C929-Gal4, and pdf-Gal4) is required and sufficient for wild-type levels of sleep maintenance. Furthermore, anti-SPR staining confirmed the SPR expression in two major subsets of pdf neurons, l-LNvs and s-LNvs. In particular, l-LNvs are common to all three Gal4 populations. Thus, the most parsimonious explanation of the results is that SPR in l-LNvs mediates a sleep-related MIP function. This rationale is also supported by previous reports. Firstly, l-LNvs respond to light and other modulatory cues and promote arousal by releasing PDF, a major wake-promoting factor functionally analogous to vertebrate orexin/hypocretin. Secondly, excitation of l-LNvs suppresses night-time sleep. Third, l-LNvs are major targets of inhibitory GABA-GABAA signaling, which promotes sleep both in flies and mammals. Fourth, blocking sNPF-mediated inhibitory input to l-LNvs impairs sleep stability particularly in night-time. Finally, MIP signaling through SPR can down-regulate cAMP levels in l-LNvs. Together, these and the genetic data provide cogent support for a role for MIP signaling in stabilizing the sleep state by modulating l-LNvs activities through the SPR (Oh, 2014).

Another group of pdf neurons, s-LNvs are critical in timing the onset of morning behavior and are the key pacemaker cells controlling the circadian locomotor rhythm. Although the precise role of s-LNvs in sleep regulation remains less clear, previous reports implicated that s-LNvs regulate sleep mainly by relaying information from l-LNvs. In response to light and modulatory substances, such as dopamine and octopamine, l-LNvs secrete PDF, which in turn elevates cAMP levels in s-LNvs by activating the PDFR. Consistent with the role of s-LNvs in sleep regulation, knockdown of PDFR in pdf neurons (presumably affecting the s-LNvs and not the l-LNvs) elevates total sleep. Recently, it was shown that s-LNvs produce sNPF, which modulates l-LNvs and stabilizes sleep, particularly in night-time. This study also reports that SPR expression in s-LNvs is important for maintaining daily sleep architecture. Knockdown of SPR in s-LNvs reduced daytime sleep and its average bout duration, whereas knockdown of SPR in l-LNvs reduced night-time sleep and its average bout duration. Together with previous results, these observations suggest that s-LNvs are involved in sleep regulation, and that MIP-SPR signaling stabilizes sleep by modulating the activity of s-LNvs directly and indirectly through l-LNvs (Oh, 2014).

The genetic and cAMP imaging results indicate that MIP regulates sleep as a ligand for SPR. Thus, it is also important to know whether MIP is secreted at biologically relevant times. The monitoring of levels of MIP peptide and mRNA at various time points in a day suggested that almost all brain MIP neurons release their contents synchronously from dusk to dawn, when the majority of flies fall and stay asleep. The rhythmic secretory activity of MIP neurons is likely to be under the control of the circadian clock rather than environmental light, because initiation and termination of the MIP secretion occurs prior to the light-off (ZT 12) and the light-on time (ZT 24), respectively. Since MIP neurons are not part of the circadian clock network (Kolodziejczyk, 2011), it would be interesting to see in the future to elucidate how they interact with the neuronal circadian clock network (Oh, 2014).

The current results indicate that MIP release in most brain neurons appears synchronized, and MIP neurons in the brain arborize in many areas of the brain, including the olfactory glomeruli, the SOG, the lateral ventral protocerebrum, mushroom body, and so on (for further evidence, see Kolodziejczyk, 2011 and Jiang, 2013). Considering SPR is expressed broadly in large numbers of the brain neurons, massively secreted MIP in these sites probably modulates not only neurons important for locomotor activities and but also many others involved in diverse biological processes, such as olfactory, feeding, sexual activity, learning, and memory (Oh, 2014).

Like in the human situation, sleep in Drosophila is also affected by other behavioral aspects, such as stress, social interactions, learning, diet, feeding, and reproduction. In females, mating suppresses daytime sleep, and male-derived SP is responsible for this sleep modulation. On the other hand, SP also plays key roles in eliciting the PMR, such as reduced receptivity to further mating and increased egg-laying. This study clearly demonstrates that the sleep-relevant SPR circuits (l-LNvs and s-LNvs) are distinct from the PMR-relevant SPR circuit (ppk+ fru+ neurons). Intriguingly, however, SP circulates in the haemolymph of mated females, raising the possibility that the haemolymph-born SP activates SPR in the sleep circuit and modulates sleep. This is certainly a plausible scenario, considering that SP is a potent agonist for the SPR, and bath-applied SPR agonist (in this case, MIP) can affect cAMP levels in s-LNvs. In theory, however, the SPR activation in the sleep circuit either by haemolymph-born SP or centrally released MIP should promote sleep, rather than suppress it. Thus, it is suspected that the daytime sleep loss observed in the mated female is not due to direct modulation of the SPR-sleep circuit by SP. Rather, SP actions on the PMR circuit elevate reproductive drives in mated females, which in consequence makes them spend more time during the day searching for food and egg-laying sites, and less time in falling asleep. Nevertheless, it cannot be formally excluded that SP modulates female sleep. In theory, SP circulating in haemolymph of the mated female can promote sleep drive and counter wakefulness driven by reproductive motivations. Thus, it is speculated that SPR may serve as a molecular integrator that computes reproductive-state coding signal (SP) and sleep-pressure coding signal (MIP) and therefore contribute to shaping daily sleep architecture (Oh, 2014).

Multiple lines of evidence indicate that MIP-SPR signaling is a part of the homeostatic control system. First, mutants lacking either MIP or SPR show significant reduction in total amount of sleep, which is an indicator of homeostatic regulation. Second, sleep deprivation drives MIP-LMIo, a subset of brain MIP neurons to release MIP into the optic lobe medulla where pdf neurons innervate. It is proposed that MIP-LMIo senses sleep pressure and modulates MIP secretion to maintain optimum sleep duration. Lastly and most importantly, mutants lacking either MIP or SPR show no sleep rebound after sleep deprivation. Together, these observations suggest that the activity of MIP neurons is controlled by two separable pathways; one associated with the circadian clock network, and the other associated with a sleep homeostat (Oh, 2014).

It has been proposed in mammals that activity-dependent metabolites, such as adenosine, GABA, prostaglandins, and cytokines, are involved in sleep homeostasis, particularly the sleep initiation phase. The role of GABA signaling in sleep is conserved both in mammals and flies. In Drosophila, GABA regulates both sleep initiation and maintenance because silencing GABAergic neurons results in a significant decrease of sleep latency from lights-off as well as mean sleep-bout duration. At the beginning of the night, GABA initiates sleep by inhibiting the activities of wake-promoting pdf neurons through the GABAA receptor, a ligand-gated Cl channel. After animals fall asleep, at least three modulatory pathways stabilize the sleep state and sustain it throughout the night: sNPF-sNPF receptor, GABA-GABAB receptor 2, and MIP-SPR. All three pathways feed inhibitory modulation into l-LNvs, and in consequence keep these neurons from releasing PDF during the night. Unlike the other two pathways, MIP-SPR signaling is also important for stabilizing daytime sleep. The model presented in this paper predicts that in the morning, shortly before light-on, s-LNvs release less sNPF than PDF. This probably is due to faster depletion of sNPF pool in s-LNvs during the night, as suggested by the fact that sNPF mRNA levels in s-LNvs are 30-fold higher in the morning (ZT 0) than in the evening (ZT 12) . Then, subsequent to light-on l-LNvs are stimulated to release PDF, which in turn modulates the motor control centers either directly or indirectly through s-LNvs, and in consequence promotes wakefulness. Later, as sleep pressure builds up during the day, MIP-LMIos sense the sleep pressure and release MIP, allowing sleep to ensue in the middle of the day. MIP is expected to act via volume transmission, meaning that once released, it can access both l-LNvs as well as s-LNvs. In daytime, however the inhibitory actions of MIP on l-LNvs are fully countered by excitatory inputs from environmental light via dopamine and octopamine signaling, partly because MIP secretion is weaker at this time of day than at night-time. For siesta sleep, therefore SPR activation in s-LNvs is more important than that in l-LNvs (Oh, 2014).

Several lines of evidence indicate that MIP, not SP, is the ancestral ligand of SPR. MIP can activate SPRs from diverse species including the sea slug Aplysia, whereas SP can only activate SPRs from Drosophila species at physiological levels. MIPs are also more potent than SP as SPR agonists. Furthermore, orthologs of SPR and MIP are clearly detectable in most (but not all) sequenced genomes from Lophotrochozoa and Ecdysozoa. By contrast, SP has been found only in the genomes of a few closely related Drosophila species, indicative of their recent origin. Together, these observations suggest that the SPR-MIP signaling axis is evolutionarily ancestral, whereas the SPR-SP signaling axis arose only recently in Drosophila evolution, concomitantly with the emergence of SP. Our discovery that sleep regulation is a possible ancestral SPR function is a critical step forward in understanding how the SPR evolved functional multiplicity by recruiting a newly emerging ligand (Oh, 2014).

The degradome and the evolution of Drosophila sex peptide as a ligand for the MIP receptor

The male sex peptide (SP) of Drosophila melanogaster has wide ranging effects on females, including rejection of courting males, increased egg production, changes to the feeding habit, increased synthesis of antimicrobial peptides and elevated locomotor activity during day-time. The peptide activates receptors in sensory neurons of the female reproductive tract and can also traverse into the hemolymph and reach the central nervous system. The SP receptor involved in rejection and egg-laying responses has been shown to be identical to the receptor for the evolutionary conserved myoinhibitory peptides (MIPs) that function as neuropeptides in both males and females. Intriguingly, MIPs cannot substitute for SP when either expressed in the male accessory glands or injected into virgin females. MIPs are linear peptides with an amidated C-terminus which protects them from cleavage by carboxypeptidases, but leaves them exposed to potential attack from aminopeptidase and endopeptidase activities. In contrast, the SP region responsible for eliciting the post-mating response is cyclic and has several hydroxyproline residues N-terminal to the disulfide bridge which is expected to protect the biological activity of SP from peptidases of the male accessory gland and seminal fluid. This study presents in vitro data showing that SP is metabolically stable, whereas MIPs are much more susceptible to degradation by peptidases of the male accessory gland and the hemolymph of virgin female D. melanogaster. SP has evolved relatively recently as a MIP receptor ligand that is particularly well adapted to surviving in the hostile degradome of the male accessory gland and seminal fluid (Isaac, 2014).

MIPs are ancestral ligands for the sex peptide receptor

Upon mating, females of many animal species undergo dramatic changes in their behavior. In Drosophila melanogaster, postmating behaviors are triggered by sex peptide (SP), which is produced in the male seminal fluid and transferred to female during copulation. SP modulates female behaviors via sex peptide receptor (SPR) located in a small subset of internal sensory neurons that innervate the female uterus and project to the CNS. Although required for postmating responses only in these female sensory neurons, SPR is expressed broadly in the CNS of both sexes. Moreover, SPR is also encoded in the genomes of insects that lack obvious SP orthologs. These observations suggest that SPR may have additional ligands and functions. This study identified myoinhibitory peptides (MIPs) as a second family of SPR ligands that is conserved across a wide range of invertebrate species. MIPs are potent agonists for Drosophila, Aedes, and Aplysia SPRs in vitro, yet are unable to trigger postmating responses in vivo. In contrast to SP, MIPs are not produced in male reproductive organs, and are not required for postmating behaviors in Drosophila females. It is concluded that MIPs are evolutionarily conserved ligands for SPR, which are likely to mediate functions other than the regulation of female reproductive behaviors (Kim, 2010).

This study presents evidence that MIPs are ligands for SPR. The evolutionary conservation of both MIPs and SPR across most of the invertebrates, including both ecdysozoa and lophotrochozoa, stands in striking contrast to the recent origin of SP, which can only be detected in the genomes of a limited number of Drosophila species. It is also noted that the sequenced hymenopteran genomes that lack SPR also lack MIPs. It is therefore concluded that MIPs are the ancestral ligands for SPR. Because MIPs were not detected in the male reproductive organs of any species tested, and MIPs are neither required nor sufficient to trigger postmating changes in Drosophila females, it is inferred that the ancestral function for MIPs and SPR is most likely unrelated to the regulation of female reproductive behavior. Genetic analysis in Drosophila has, however, not yet revealed any other functions for SPR that might be ascribed to MIPs. This may be due to functional redundancy with other putative MIP receptors, such as CG30106 and its closely related homolog CG14593. In other insects, MIPs have been shown to suppress visceral muscle contractions, regulate production of juvenile hormones and ecdysteroid, and modulate ecdysis behavior. It remains to be seen whether any of these functions are mediated by SPR (Kim, 2010).

Nonetheless, the data do not formally exclude the possibility that SPR regulates female reproductive behavior in species other than Drosophila. This study also found that Aedes contains at least one other potent SPR ligand, presumably unrelated to both MIPs and SP. It is possible that this ligand may function in Aedes in a manner analogous to SP in Drosophila (Kim, 2010).

A novel wide-field neuron with branches in the lamina of the Drosophila visual system expresses myoinhibitory peptide and may be associated with the clock

Although neuropeptides are widespread throughout the central nervous system of the fruifly Drosophila, no records exist of peptidergic neurons in the first synaptic region of the visual system, the lamina. This paper describes a novel type of neuron that has wide-field tangential arborizations just distal to the lamina neuropil and that expresses myoinhibitory peptide (MIP). The cell bodies of these neurons, designated lateral MIP-immunoreactive optic lobe (LMIo) neurons, lie anteriorly at the base of the medulla of the optic lobe. The LMIo neurons also arborize in several layers of the medulla and in the dorso-lateral and lateral protocerebrum. Since the LMIo resemble LN(v) clock neurons, this study has investigated the relationships between these two sets of neurons by combining MIP-immunolabeling with markers for two of the clock genes, namely, Cryptochrome and Timeless, or with antisera to two peptides expressed in clock neurons, pigment-dispersing factor and ion transport peptide. LMIo neurons do not co-express any of these clock neuron markers. However, branches of LMIo and clock neurons overlap in several regions. Furthermore, the varicose lamina branches of LMIo neurons superimpose those of two large bilateral serotonergic neurons. The close apposition of the terminations of MIP- and serotonin-producing neurons distal to the lamina suggests that they have the same peripheral targets. These data indicate that the LMIo neurons are not bona fide clock neurons, but they may be associated with the clock system and regulate signaling peripherally in the visual system (Kolodziejczyk, 2010).

Myoinhibiting peptides are the ancestral ligands of the promiscuous Drosophila sex peptide receptor

Male insects change behaviors of female partners by co-transferring accessory gland proteins (Acps) like sex peptide (SP), with their sperm. The Drosophila sex peptide receptor (SPR) is a G protein-coupled receptor expressed in the female's nervous system and genital tract. While most Acps show a fast rate of evolution, SPRs are highly conserved in insects. This study reports activation of SPRs by evolutionary conserved myoinhibiting peptides (MIPs). Structural determinants in SP and MIPs responsible for this dual receptor activation are characterized. Drosophila SPR is also expressed in embryonic and larval stages and in the adult male nervous system, whereas SP expression is restricted to the male reproductive system. MIP transcripts occur in male and female central nervous system, possibly acting as endogenous SPR ligands. Evolutionary consequences of the promiscuous nature of SPRs are discussed. MIPs likely function as ancestral ligands of SPRs and could place evolutionary constraints on the MIP/SPR class (Poels, 2010).


Search PubMed for articles about Drosophila Mip

Carlsson, M. A., Diesner, M., Schachtner, J. and Nassel, D. R. (2010). Multiple neuropeptides in the Drosophila antennal lobe suggest complex modulatory circuits. J Comp Neurol 518: 3359-3380. PubMed ID: 20575072

Conzelmann, M., Williams, E. A., Tunaru, S., Randel, N., Shahidi, R., Asadulina, A., Berger, J., Offermanns, S. and Jekely, G. (2013). Conserved MIP receptor-ligand pair regulates Platynereis larval settlement. Proc Natl Acad Sci U S A 110: 8224-8229. PubMed ID: 23569279

Dey, S., Chamero, P., Pru, J. K., Chien, M. S., Ibarra-Soria, X., Spencer, K. R., Logan, D. W., Matsunami, H., Peluso, J. J. and Stowers, L. (2015). Cyclic Regulation of Sensory Perception by a Female Hormone Alters Behavior. Cell 161: 1334-1344. PubMed ID: 26046438

Gou, B., Liu, Y., Guntur, A. R., Stern, U. and Yang, C. H. (2014). Mechanosensitive neurons on the internal reproductive tract contribute to egg-laying-induced acetic acid attraction in Drosophila. Cell Rep 9: 522-530. PubMed ID: 25373900

Dyer, J. O., Dutta, A., Gogol, M., Weake, V. M., Dialynas, G., Wu, X., Seidel, C., Zhang, Y., Florens, L., Washburn, M. P., Abmayr, S. M. and Workman, J. L. (2017). Myeloid Leukemia Factor acts in a chaperone complex to regulate transcription factor stability and gene expression. J Mol Biol 429(13): 2093-2107. PubMed ID: 27984043

Hasemeyer, M., Yapici, N., Heberlein, U. and Dickson, B. J. (2009). Sensory neurons in the Drosophila genital tract regulate female reproductive behavior. Neuron 61: 511-518. PubMed ID: 19249272

Hussain, A., Zhang, M., Ucpunar, H. K., Svensson, T., Quillery, E., Gompel, N., Ignell, R. and Grunwald Kadow, I. C. (2016a). Ionotropic chemosensory receptors mediate the taste and smell of polyamines. PLoS Biol 14: e1002454. PubMed ID: 27145030

Hussain, A., Ucpunar, H. K., Zhang, M., Loschek, L. F. and Grunwald Kadow, I. C. (2016b). Neuropeptides modulate female chemosensory processing upon mating in Drosophila. PLoS Biol 14: e1002455. PubMed ID: 27145127

Hull, J. J. and Brent, C. S. (2014). Identification and characterization of a sex peptide receptor-like transcript from the western tarnished plant bug Lygus hesperus. Insect Mol Biol 23: 301-319. PubMed ID: 24467643

Isaac, R. E., Kim, Y. J. and Audsley, N. (2014). The degradome and the evolution of Drosophila sex peptide as a ligand for the MIP receptor. Peptides 53: 258-264. PubMed ID: 24398368

Jiang, H., Lkhagva, A., Daubnerova, I., Chae, H. S., Simo, L., Jung, S. H., Yoon, Y. K., Lee, N. R., Seong, J. Y., Zitnan, D., Park, Y. and Kim, Y. J. (2013). Natalisin, a tachykinin-like signaling system, regulates sexual activity and fecundity in insects. Proc Natl Acad Sci U S A 110: E3526-3534. PubMed ID: 23980168

Kim, Y. J., Bartalska, K., Audsley, N., Yamanaka, N., Yapici, N., Lee, J. Y., Kim, Y. C., Markovic, M., Isaac, E., Tanaka, Y. and Dickson, B. J. (2010). MIPs are ancestral ligands for the sex peptide receptor. Proc Natl Acad Sci U S A 107: 6520-6525. PubMed ID: 20308537

Kolodziejczyk, A. and Nassel, D. R. (2011). A novel wide-field neuron with branches in the lamina of the Drosophila visual system expresses myoinhibitory peptide and may be associated with the clock. Cell Tissue Res 343: 357-369. PubMed ID: 21174124

Kubli, E. (2003). Sex-peptides: seminal peptides of the Drosophila male. Cell Mol Life Sci 60: 1689-1704. PubMed ID: 14504657

Lefèvre, P. L., Palin, M. F. and Murphy, B. D. (2011). Polyamines on the reproductive landscape. Endocr Rev 32: 694-712. PubMed ID: 21791568

Miller, M., Chen, A., Gobert, V., Auge, B., Beau, M., Burlet-Schiltz, O., Haenlin, M. and Waltzer, L. (2017). Control of RUNX-induced repression of Notch signaling by MLF and its partner DnaJ-1 during Drosophila hematopoiesis. PLoS Genet 13(7): e1006932. PubMed ID: 28742844

Oh, Y., Yoon, S. E., Zhang, Q., Chae, H. S., Daubnerova, I., Shafer, O. T., Choe, J. and Kim, Y. J. (2014). A homeostatic sleep-stabilizing pathway in Drosophila composed of the sex peptide receptor and its ligand, the myoinhibitory peptide. PLoS Biol 12: e1001974. PubMed ID: 25333796

Poels, J., Van Loy, T., Vandersmissen, H. P., Van Hiel, B., Van Soest, S., Nachman, R. J. and Vanden Broeck, J. (2010). Myoinhibiting peptides are the ancestral ligands of the promiscuous Drosophila sex peptide receptor. Cell Mol Life Sci 67: 3511-3522. PubMed ID: 20458515

Root, C. M., Ko, K. I., Jafari, A. and Wang, J. W. (2011). Presynaptic facilitation by neuropeptide signaling mediates odor-driven food search. Cell 145: 133-144. PubMed ID: 21458672

Siju, K. P., Reifenrath, A., Scheiblich, H., Neupert, S., Predel, R., Hansson, B. S., Schachtner, J. and Ignell, R. (2014). Neuropeptides in the antennal lobe of the yellow fever mosquito, Aedes aegypti. J Comp Neurol 522: 592-608. PubMed ID: 23897410

Silbering, A. F., Rytz, R., Grosjean, Y., Abuin, L., Ramdya, P., Jefferis, G. S. and Benton, R. (2011). Complementary function and integrated wiring of the evolutionarily distinct Drosophila olfactory subsystems. J Neurosci 31: 13357-13375. PubMed ID: 21940430

Walker, S. J., Corrales-Carvajal, V. M. and Ribeiro, C. (2015). Postmating Circuitry Modulates Salt Taste Processing to Increase Reproductive Output in Drosophila. Curr Biol 25: 2621-2630. PubMed ID: 26412135

Yang, C. H., Rumpf, S., Xiang, Y., Gordon, M. D., Song, W., Jan, L. Y. and Jan, Y. N. (2009). Control of the postmating behavioral switch in Drosophila females by internal sensory neurons. Neuron 61: 519-526. PubMed ID: 19249273

Biological Overview

date revised: 5 November 2023

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