Marie-Laure Parmentier

The Partner of Inscuteable/Discs-Large Complex Is Required to Establish Planar Polarity during Asymmetric Cell Division in Drosophila

Frizzled (Fz) signaling regulates cell polarity in both vertebrates and invertebrates. In Drosophila, Fz orients the asymmetric division of the sensory organ precursor cell (pI) along the antero-posterior axis of the notum. Planar polarization involves a remodeling of the apical-basal polarity of the pI cell. The Discs-large (Dlg) and Partner of Inscuteable (Pins) proteins accumulate at the anterior cortex, while Bazooka (Baz) relocalizes to the posterior cortex. Dlg interacts directly with Pins and regulates the localization of Pins and Baz. Pins acts with Fz to localize Baz posteriorly, but Baz is not required to localize Pins anteriorly. Finally, Baz and the Dlg/Pins complex are required for the asymmetric localization of Numb. Thus, the Dlg/Pins complex responds to Fz signaling to establish planar asymmetry in the pI cell.

Muscle dystroglycan organizes the postsynapse and regulates presynaptic neurotransmitter release at the Drosophila neuromuscular junction.

Background The Dystrophin-glycoprotein complex (DGC) comprises dystrophin, dystroglycan, sarcoglycan, dystrobrevin and syntrophin subunits. In muscle fibers, it is thought to provide an essential mechanical link between the intracellular cytoskeleton and the extracellular matrix and to protect the sarcolemma during muscle contraction. Mutations affecting the DGC cause muscular dystrophies. Most members of the DGC are also concentrated at the neuromuscular junction (NMJ), where their deficiency is often associated with NMJ structural defects. Hence, synaptic dysfunction may also intervene in the pathology of dystrophic muscles. Dystroglycan is a central component of the DGC because it establishes a link between the extracellular matrix and Dystrophin. In this study, we focused on the synaptic role of Dystroglycan (Dg) in Drosophila. Methodology/Principal Findings We show that Dg was concentrated postsynaptically at the glutamatergic NMJ, where, like in vertebrates, it controls the concentration of synaptic Laminin and Dystrophin homologues. We also found that synaptic Dg controlled the amount of postsynaptic 4.1 protein Coracle and alpha-Spectrin, as well as the relative subunit composition of glutamate receptors. In addition, both Dystrophin and Coracle were required for normal Dg concentration at the synapse. In electrophysiological recordings, loss of postsynaptic Dg did not affect postsynaptic response, but, surprisingly, led to a decrease in glutamate release from the presynaptic site. Conclusion/Significance Altogether, our study illustrates a conservation of DGC composition and interactions between Drosophila and vertebrates at the synapse, highlights new proteins associated with this complex and suggests an unsuspected trans-synaptic function of Dg.

Protective role of Engrailed in a Drosophila model of Huntington's disease

Huntingtons disease (HD) is caused by the expansion of the polyglutamine (polyQ) tract in the human Huntingtin (hHtt) protein (polyQ-hHtt). Although this mutation behaves dominantly, htt loss of function may also contribute to HD pathogenesis. Using a Drosophila model of HD, we found that Engrailed (EN), a transcriptional activator of endogenous Drosophila htt (dhtt), is able to prevent aggregation of polyQ-hHtt. To interpret these findings, we tested and identified a protective role of N-terminal fragments of both Drosophila and Human wild-type Htt onto polyQ-hHtt-induced cellular defects. In addition, N-terminal parts of normal hHtt were also able to rescue eye degeneration due to the loss of Drosophila endogenous dhtt function. Thus, our data indicate that Drosophila and Human Htt share biological properties, and confirm a model whereby EN activates endogenous dhtt, which in turn prevents polyQ-hHtt-induced phenotypes. The protective role of wild-type hHtt N-terminal parts, specifically onto polyQ-hHtt-induced cellular toxicity suggests that the HD may be considered as a dominant negative disease rather than solely dominant.

A Presynaptic Role of Microtubule-Associated Protein 1/Futsch in Drosophila: Regulation of Active Zone Number and Neurotransmitter Release

Structural microtubule-associated proteins (MAPs), like MAP1, not only control the stability of microtubules, but also interact with postsynaptic proteins in the nervous system. Their presynaptic role has barely been studied. To tackle this question, we used the Drosophila model in which there is only one MAP1 homolog: Futsch, which is expressed at the larval neuromuscular junction, presynaptically only. We show that Futsch regulates neurotransmitter release and active zone density. Importantly, we provide evidence that this role of Futsch is not just the consequence of its microtubule-stabilizing function. Using high-resolution microscopy, we show that Futsch and microtubules are almost systematically present in close proximity to active zones, with Futsch being localized in-between microtubules and active zones. Using proximity ligation assays, we further demonstrate the proximity of Futsch, but not microtubules, to active zone components. Altogether our data are in favor of a model by which Futsch locally stabilizes active zones, by reinforcing their link with the underlying microtubule cytoskeleton.

A huntingtin peptide inhibits polyQ-huntingtin associated defects.

Background Huntington’s disease (HD) is caused by the abnormal expansion of the polyglutamine tract in the human Huntingtin protein (polyQ-hHtt). Although this mutation behaves dominantly, huntingtin loss of function also contributes to HD pathogenesis. Indeed, wild-type Huntingtin plays a protective role with respect to polyQ-hHtt induced defects. Methodology/Principal Findings The question that we addressed here is what part of the wild-type Huntingtin is responsible for these protective properties. We first screened peptides from the Huntingtin protein in HeLa cells and identified a 23 aa peptide (P42) that inhibits polyQ-hHtt aggregation. P42 is part of the endogenous Huntingtin protein and lies within a region rich in proteolytic sites that plays a critical role in the pathogenesis process. Using a Drosophila model of HD, we tested the protective properties of this peptide on aggregation, as well as on different polyQ-hHtt induced neuronal phenotypes: eye degeneration (an indicator of cell death), impairment of vesicular axonal trafficking, and physiological behaviors such as larval locomotion and adult survival. Together, our results demonstrate high protective properties for P42 in vivo, in whole animals. These data also demonstrate a specific role of P42 on Huntington’s disease model, since it has no effect on other models of polyQ-induced diseases, such as spinocerebellar ataxias. Conclusions/Significance Altogether our data show that P42, a 23 aa-long hHtt peptide, plays a protective role with respect to polyQ-hHtt aggregation as well as cellular and behavioral dysfunctions induced by polyQ-hHtt in vivo. Our study also confirms the correlation between polyQ-hHtt aggregation and neuronal defects. Finally, these results strongly suggest a therapeutic potential for P42, specific of Huntington’s disease.

Tau excess impairs mitosis and kinesin-5 function, leading to aneuploidy and cell death.

ABSTRACT In neurodegenerative diseases such as Alzheimers disease (AD), cell cycle defects and associated aneuploidy have been described. However, the importance of these defects in the physiopathology of AD and the underlying mechanistic processes are largely unknown, in particular with respect to the microtubule (MT)-binding protein Tau, which is found in excess in the brain and cerebrospinal fluid of affected individuals. Although it has long been known that Tau is phosphorylated during mitosis to generate a lower affinity for MTs, there is, to our knowledge, no indication that an excess of this protein could affect mitosis. Here, we studied the effect of an excess of human Tau (hTau) protein on cell mitosis in vivo. Using the Drosophila developing wing disc epithelium as a model, we show that an excess of hTau induces a mitotic arrest, with the presence of monopolar spindles. This mitotic defect leads to aneuploidy and apoptotic cell death. We studied the mechanism of action of hTau and found that the MT-binding domain of hTau is responsible for these defects. We also demonstrate that the effects of hTau occur via the inhibition of the function of the kinesin Klp61F, the Drosophila homologue of kinesin-5 (also called Eg5 or KIF11). We finally show that this deleterious effect of hTau is also found in other Drosophila cell types (neuroblasts) and tissues (the developing eye disc), as well as in human HeLa cells. By demonstrating that MT-bound Tau inhibits the Eg5 kinesin and cell mitosis, our work provides a new framework to consider the role of Tau in neurodegenerative diseases. Drosophila Collection: We show that Tau, a microtubule-binding protein involved in many neurodegenerative diseases, impairs mitosis when in excess. We show that this occurs via the inhibition of the kinesin-5 mitotic motor.

Honey Bee Allatostatins Target Galanin/Somatostatin-Like Receptors and Modulate Learning: A Conserved Function?

Sequencing of the honeybee genome revealed many neuropeptides and putative neuropeptide receptors, yet functional characterization of these peptidic systems is scarce. In this study, we focus on allatostatins, which were first identified as inhibitors of juvenile hormone synthesis, but whose role in the adult honey bee (Apis mellifera) brain remains to be determined. We characterize the bee allatostatin system, represented by two families: allatostatin A (Apime-ASTA) and its receptor (Apime-ASTA-R); and C-type allatostatins (Apime-ASTC and Apime-ASTCC) and their common receptor (Apime-ASTC-R). Apime-ASTA-R and Apime-ASTC-R are the receptors in bees most closely related to vertebrate galanin and somatostatin receptors, respectively. We examine the functional properties of the two honeybee receptors and show that they are transcriptionally expressed in the adult brain, including in brain centers known to be important for learning and memory processes. Thus we investigated the effects of exogenously applied allatostatins on appetitive olfactory learning in the bee. Our results show that allatostatins modulate learning in this insect, and provide important insights into the evolution of somatostatin/allatostatin signaling.

Drosophila Nesprin-1 controls glutamate receptor density at neuromuscular junctions

Nesprin-1 is a core component of a protein complex connecting nuclei to cytoskeleton termed LINC (linker of nucleoskeleton and cytoskeleton). Nesprin-1 is anchored to the nuclear envelope by its C-terminal KASH domain, the disruption of which has been associated with neuronal and neuromuscular pathologies, including autosomal recessive cerebellar ataxia and Emery–Dreifuss muscular dystrophy. Here, we describe a new and unexpected role of Drosophila Nesprin-1, Msp-300, in neuromuscular junction. We show that larvae carrying a deletion of Msp-300 KASH domain (Msp-300∆KASH) present a locomotion defect suggestive of a myasthenia, and demonstrate the importance of muscle Msp-300 for this phenotype, using tissue-specific RNAi knock-down. We show that Msp-300∆KASH mutants display abnormal neurotransmission at the larval neuromuscular junction, as well as an imbalance in postsynaptic glutamate receptor composition with a decreased percentage of GluRIIA-containing receptors. We could rescue Msp-300∆KASH locomotion phenotypes by GluRIIA overexpression, suggesting that the locomotion impairment associated with the KASH domain deletion is due to a reduction in junctional GluRIIA. In summary, we found that Msp-300 controls GluRIIA density at the neuromuscular junction. Our results suggest that Drosophila is a valuable model for further deciphering how Nesprin-1 and LINC disruption may lead to neuronal and neuromuscular pathologies.

Mushroom body neuronal remodelling is necessary for short-term but not for long-term courtship memory in Drosophila.

The remodelling of neurons during their development is considered necessary for their normal function. One fundamental mechanism involved in this remodelling process in both vertebrates and invertebrates is axon pruning. A well‐documented case of such neuronal remodelling is the developmental axon pruning of mushroom body γ neurons that occurs during metamorphosis in Drosophila. The γ neurons undergo pruning of larval‐specific dendrites and axons at metamorphosis, followed by their regrowth as adult‐specific dendrites and axons. We recently revealed a molecular cascade required for this pruning. The nuclear receptor ftz‐f1 activates the expression of the steroid hormone receptor EcR‐B1, a key component for γ remodelling, and represses expression of Hr39, an ftz‐f1 homologous gene. If ectopically expressed in the γ neurons, HR39 inhibits normal pruning, probably by competing with endogenous FTZ‐F1, which results in decreased EcR‐B1 expression. The mushroom bodies are a bilaterally symmetric structure in the larval and adult brain and are involved in the processing of different types of olfactory memory. How memory is affected in pruning‐deficient adult flies that possess larval‐stage neuronal circuitry will help to explain the functional role of neuron remodelling. Flies overexpressing Hr39 are viable as adults and make it possible to assess the requirement for wild‐type mushroom body pruning in memory. While blocking mushroom body neuron remodelling impaired memory after short‐term courtship conditioning, long‐term memory was normal. These results show that larval pruning is necessary for adult memory and that expression of courtship short‐term memory and long‐term memory may be parallel and independent.

A New Behavioral Test and Associated Genetic Tools Highlight the Function of Ventral Abdominal Muscles in Adult Drosophila

The function of the nervous system in complex animals is reflected by the achievement of specific behaviors. For years in Drosophila, both simple and complex behaviors have been studied and their genetic bases have emerged. The neuromuscular junction is maybe one of the prototypal simplest examples. A motor neuron establishes synaptic connections on its muscle cell target and elicits behavior: the muscle contraction. Different muscles in adult fly are related to specific behaviors. For example, the thoracic muscles are associated with flight and the leg muscles are associated with locomotion. However, specific tools are still lacking for the study of cellular physiology in distinct motor neuron subpopulations. Here we decided to use the abdominal muscles and in particular the ventral abdominal muscles (VAMs) in adult Drosophila as new model to link a precise behavior to specific motor neurons. Hence, we developed a new behavioral test based on the folding movement of the adult abdomen. Further, we performed a genetic screen and identify two specific Gal4 lines with restricted expression patterns to the adult motor neurons innervating the VAMs or their precursor cells. Using these genetic tools, we showed that the lack of the VAMs or the loss of the synaptic transmission in their innervating motor neurons lead to a significant impairment of the abdomen folding behavior. Altogether, our results allow establishing a direct link between specific motor neurons and muscles for the realization of particular behavior: the folding behavior of the abdomen in Drosophila.