Drosophila Inhibitor Of Apoptosis Protein Research Articles

MiR-927 regulates insect wing development by targeting the Hippo pathway.

How organ size is determined is a fundamental question in life sciences. Recent studies have highlighted the importance of the Hippo pathway in regulating organ size. This pathway controls cell proliferation and cell death to maintain the proper number of cells. The activity of the Hippo pathway is tightly fine-tuned through various post-translational modifications, such as phosphorylation and ubiquitination. Here, we discover that miR-927 is a novel regulator of wing size. Overexpression of miR-927 decreases wing size, which can be rescued by co-expressing miR-927-sponge. Next, we show that miR-927 stimulates apoptosis and suppresses the expression of Drosophila inhibitor of apoptosis protein 1, a well-known target gene of the Hippo pathway. Genetic epistatic analyses position miR-927 upstream of Yorkie (Yki) to modulate the Hippo pathway. In addition, there is a matching miR-927 seed site in the yki 3' untranslated region (3'-UTR), and we demonstrate that yki 3'-UTR is the direct target of miR-927. Ultimately, our study reveals that the targeting of yki by miR-927 to regulate the Hippo pathway is conserved in Helicoverpa armigera. Administration of miR-927 via star polycation (SPc) nanocarrier effectively inhibits wing development in H. armigera. Taken together, our findings uncover a novel mechanism by which Yki is silenced at the post-transcriptional level by miR-927, and provide a new perspective on pest management.

Expanded polyQ aggregates interact with sarco-endoplasmic reticulum calcium ATPase and Drosophila inhibitor of apoptosis protein1 to regulate polyQ mediated neurodegeneration in Drosophila

Polyglutamine (polyQ) induced neurodegeneration is one of the leading causes of progressive neurodegenerative disorders characterized clinically by deteriorating movement defects, psychiatric disability, and dementia. Calcium [Ca2+] homeostasis, which is essential for the functioning of neuronal cells, is disrupted under these pathological conditions. In this paper, we simulated Huntington's disease phenotype in the neuronal cells of the Drosophila eye and identified [Ca2+] pump, sarco-endoplasmic reticulum calcium ATPase (SERCA), as one of the genetic modifiers of the neurodegenerative phenotype. This paper shows genetic and molecular interaction between polyglutamine (polyQ) aggregates, SERCA and DIAP1. We present evidence that polyQ aggregates interact with SERCA and alter its dynamics, resulting in a decrease in cytosolic [Ca2+] and an increase in ER [Ca2+], and thus toxicity. Downregulating SERCA lowers the enhanced calcium levels in the ER and rescues, morphological and functional defects caused due to expanded polyQ repeats. Cell proliferation markers such as Yorkie (Yki), Scalloped (Sd), and phosphatidylinositol 3 kinases/protein kinase B (PI3K/Akt), also respond to varying levels of calcium due to genetic manipulations, adding to the amelioration of degeneration. These results imply that neurodegeneration due to expanded polyQ repeats is sensitive to SERCA activity, and its manipulation can be an important step toward its therapeutic measures.

Wg/Wnt1 and Erasp link ER stress to proapoptotic signaling in an autosomal dominant retinitis pigmentosa model

The endoplasmic reticulum (ER) is a subcellular organelle essential for cellular homeostasis. Perturbation of ER functions due to various conditions can induce apoptosis. Chronic ER stress has been implicated in a wide range of diseases, including autosomal dominant retinitis pigmentosa (ADRP), which is characterized by age-dependent retinal degeneration caused by mutant rhodopsin alleles. However, the signaling pathways that mediate apoptosis in response to ER stress remain poorly understood. In this study, we performed an unbiased in vivo RNAi screen with a Drosophila ADRP model and found that Wg/Wnt1 mediated apoptosis. Subsequent transcriptome analysis revealed that ER stress-associated serine protease (Erasp), which has been predicted to show serine-type endopeptidase activity, was a downstream target of Wg/Wnt1 during ER stress. Furthermore, knocking down Erasp via RNAi suppressed apoptosis induced by mutant rhodopsin-1 (Rh-1P37H) toxicity, alleviating retinal degeneration in the Drosophila ADRP model. In contrast, overexpression of Erasp resulted in enhanced caspase activity in Drosophila S2 cells treated with apoptotic inducers and the stabilization of the initiator caspase Dronc (Death regulator Nedd2-like caspase) by stimulating DIAP1 (Drosophila inhibitor of apoptosis protein 1) degradation. These findings helped identify a novel cell death signaling pathway involved in retinal degeneration in an autosomal dominant retinitis pigmentosa model.

Apoptosis inhibition mitigates aging effects in Drosophila melanogaster.

Aging is a natural biological process that results in progressive loss of cell, tissue, and organ function. One of the causing factors of the aging process is the decrease in muscle mass, which has not been fully verified in Drosophila. Apoptotic cell death may result in aberrant cell loss and can eventually diminish tissue function and muscle atrophy. If so, inhibition of apoptosis may prolong longevity and reduce motor function and muscle mass decline with age in Drosophila flies. Here, we used Drosophila melanogaster as study material, and induced the overexpression of Drosophila inhibitor of apoptosis protein 1 gene to inhibit apoptosis, and investigated theeffect of apoptosis inhibition on the longevity and age-related declines in flight and climbing ability and muscle mass. As a result, the inhibition of apoptosis tended to mitigate the aging effects and prolonged longevity and reduced climbing ability decline with age. The current study suggests that apoptosis inhibition could mitigate the aging effects in D. melanogaster. Although such effects have already been known in mammals, the current results suggest that the apoptosis may play a similar role in insects as well.

Insecticidal effects of dsRNA targeting the Diap1 gene in dipteran pests

The Drosophila melanogaster (fruit fly) gene Diap1 encodes a protein referred to as DIAP1 (DrosophilaInhibitor of Apoptosis Protein 1) that acts to supress apoptosis in “normal” cells in the fly. In this study we investigate the use of RNA interference (RNAi) to control two dipteran pests, Musca domestica and Delia radicum, by disrupting the control of apoptosis. Larval injections of 125–500 ng of Diap1 dsRNA resulted in dose-dependent mortality which was shown to be attributable to down-regulation of target mRNA. Insects injected with Diap1 dsRNA have approx. 1.5-2-fold higher levels of caspase activity than controls 24 hours post injection, providing biochemical evidence that inhibition of apoptotic activity by the Diap1 gene product has been decreased. By contrast adults were insensitive to injected dsRNA. Oral delivery failed to induce RNAi effects and we suggest this is attributable to degradation of ingested dsRNA by intra and extracellular RNAses. Non-target effects were demonstrated via mortality and down-regulation of Diap1 mRNA levels in M. domestica larvae injected with D. radicum Diap1 dsRNA, despite the absence of 21 bp identical sequence regions in the dsRNA. Here we show that identical 15 bp regions in dsRNA are sufficient to trigger non-target RNAi effects.

Drosophila SETDB1 and caspase cooperatively fine-tune cell fate determination of sensory organ precursor.

Drosophila produce a constant number of mechanosensory bristles called macrochaetae (MC), which develop from sensory organ precursor (SOP) cells within a proneural cluster (PNC). However, what ensures the precise determination of SOP cells remains to be elucidated. In this study, we conducted RNAi screening in PNC for genes involved in epigenetic regulation. We identified a H3K9 histone methyltransferase, SETDB1/eggless, as a regulator of SOP development. Knockdown of SETDB1 in PNC led to additional SOPs. We further tested the relationship between SETDB1 and non-apoptotic function of caspase on SOP development. Reinforcing caspase activation by heterozygous Drosophila inhibitor of apoptosis protein 1 (DIAP1) mutation rescued ectopic SOP development caused by SETDB1 knockdown. Knockdown of SETDB1, however, had little effect on caspase activity. Simultaneous loss of SETDB1 and caspase activity resulted in further increase in MC, indicating that the two components work cooperatively. Our study suggests the fine-tuning mechanisms for SOP development by epigenetic methyltransferase and non-apoptotic caspase function.

Ubr3 E3 ligase regulates apoptosis by controlling the activity of DIAP1 in Drosophila.

Apoptosis has essential roles in a variety of cellular and developmental processes. Although the pathway is well studied, how the activities of individual components in the pathway are regulated is less understood. In Drosophila, a key component in apoptosis is Drosophila inhibitor of apoptosis protein 1 (DIAP1), which is required to prevent caspase activation. Here, we demonstrate that Drosophila CG42593 (ubr3), encoding the homolog of mammalian UBR3, has an essential role in regulating the apoptosis pathway. We show that loss of ubr3 activity causes caspase-dependent apoptosis in Drosophila eye and wing discs. Our genetic epistasis analyses show that the apoptosis induced by loss of ubr3 can be suppressed by loss of initiator caspase Drosophila Nedd2-like caspase (Dronc), or by ectopic expression of the apoptosis inhibitor p35, but cannot be rescued by overexpression of DIAP1. Importantly, we show that the activity of Ubr3 in the apoptosis pathway is not dependent on its Ring-domain, which is required for its E3 ligase activity. Furthermore, we find that through the UBR-box domain, Ubr3 physically interacts with the neo-epitope of DIAP1 that is exposed after caspase-mediated cleavage. This interaction promotes the recruitment and ubiquitination of substrate caspases by DIAP1. Together, our data indicate that Ubr3 interacts with DIAP1 and positively regulates DIAP1 activity, possibly by maintaining its active conformation in the apoptosis pathway.

Gga-miR-375 Plays a Key Role in Tumorigenesis Post Subgroup J Avian Leukosis Virus Infection

Avian leukosis is a neoplastic disease caused in part by subgroup J avian leukosis virus J (ALV-J). Micro ribonucleic acids (miRNAs) play pivotal oncogenic and tumour-suppressor roles in tumour development and progression. However, little is known about the potential role of miRNAs in avian leukosis tumours. We have found a novel tumour-suppressor miRNA, gga-miR-375, associated with avian leukosis tumorigenesis by miRNA microarray in a previous report. We have also previously studied the biological function of gga-miR-375; Overexpression of gga-miR-375 significantly inhibited DF-1 cell proliferation, and significantly reduced the expression of yes-associated protein 1 (YAP1) by repressing the activity of a luciferase reporter carrying the 3′-untranslated region of YAP1. This indicates that gga-miR-375 is frequently downregulated in avian leukosis by inhibiting cell proliferation through YAP1 oncogene targeting. Overexpression of gga-miR-375 markedly promoted serum starvation induced apoptosis, and there may be the reason why the tumour cycle is so long in the infected chickens. In vivo assays, gga-miR-375 was significantly downregulated in chicken livers 20 days after infection with ALV-J, and YAP1 was significantly upregulated 20 days after ALV-J infection (P<0.05). We also found that expression of cyclin E, an important regulator of cell cycle progression, was significantly upregulated (P<0.05). Drosophila inhibitor of apoptosis protein 1 (DIAP1), which is related to caspase-dependent apoptosis, was also significantly upregulated after infection. Our data suggests that gga-miR-375 may function as a tumour suppressor thereby regulating cancer cell proliferation and it plays a key role in avian leukosis tumorigenesis.

Non-cell autonomous control of apoptosis by ligand-independent Hedgehog signaling in Drosophila

Hedgehog (Hh) signaling is important for development and homeostasis in vertebrates and invertebrates. Ligand-independent, deregulated Hh signaling caused by loss of negative regulators such as Patched causes excessive cell proliferation, leading to overgrowth in Drosophila and tumors in humans, including basal-cell carcinoma and medulloblastoma. We show that in Drosophila deregulated Hh signaling also promotes cell survival by increasing the resistance to apoptosis. Surprisingly, cells with deregulated Hh activity do not protect themselves from apoptosis; instead, they promote cell survival of neighboring wild-type cells. This non-cell autonomous effect is mediated by Hh-induced Notch signaling, which elevates the protein levels of Drosophila inhibitor of apoptosis protein-1 (Diap-1), conferring resistance to apoptosis. In summary, we demonstrate that deregulated Hh signaling not only promotes proliferation but also cell survival of neighboring cells. This non-cell autonomous control of apoptosis highlights an underappreciated function of deregulated Hh signaling, which may help to generate a supportive micro-environment for tumor development.

Notch Signaling Activates Yorkie Non-Cell Autonomously in Drosophila

In Drosophila imaginal epithelia, cells mutant for the endocytic neoplastic tumor suppressor gene vps25 stimulate nearby untransformed cells to express Drosophila Inhibitor-of-Apoptosis-Protein-1 (DIAP-1), conferring resistance to apoptosis non-cell autonomously. Here, we show that the non-cell autonomous induction of DIAP-1 is mediated by Yorkie, the conserved downstream effector of Hippo signaling. The non-cell autonomous induction of Yorkie is due to Notch signaling from vps25 mutant cells. Moreover, activated Notch in normal cells is sufficient to induce non-cell autonomous Yorkie activity in wing imaginal discs. Our data identify a novel mechanism by which Notch promotes cell survival non-cell autonomously and by which neoplastic tumor cells generate a supportive microenvironment for tumor growth.

Screening of binding proteins that interact with human Salvador 1 in a human fetal liver cDNA library by the yeast two-hybrid system

Salvador promotes both cell cycle exit and apoptosis through the modulation of both cyclin E and Drosophila inhibitor of apoptosis protein in Drosophila. However, the cellular function of human Salvador (hSav1) is rarely reported. To screen for novel binding proteins that interact with hSav1, the cDNA of hSav1 was cloned into a bait protein plasmid, and positive clones were screened from a human fetal liver cDNA library by the yeast two-hybrid system. hSav1 mRNA was expressed in yeast and there was no self-activation and toxicity in the yeast strain AH109. Twenty proteins were found to interact with hSav1, including HS1 (haematopoietic cell specific protein1)-associated protein X-1 (HAX-1); neural precursor cell expressed, developmentally down-regulated 9, pyruvate kinase, liver and RBC, cytochrome c oxidase subunit Vb, enoyl coenzyme A hydratase short chain 1, and NADH dehydrogenase (ubiquinone) 1 beta subcomplex, demonstrating that the yeast two-hybrid system is an efficient method for investigating protein interactions. Among the identified proteins, there were many mitochondrial proteins, indicating that hSav1 may play a role in mitochondrial function. We also confirmed the interaction of HAX-1 and hSav1 in mammalian cells. This investigation provides functional clues for further exploration of potential apoptosis-related proteins in disease biotherapy.

The NAB-Brk Signal Bifurcates at JNK to Independently Induce Apoptosis and Compensatory Proliferation

Apoptosis operates to eliminate damaged or potentially dangerous cells. This loss is often compensated by extra proliferation of neighboring cells. Studies in Drosophila imaginal discs suggest that the signal for the additional growth emanates from the dying cells. In particular, it was suggested that the initiator caspase Dronc mediates compensatory proliferation (CP) through Dp53 in wing discs. However, the exact mechanism that governs this CP remained poorly understood. We have previously shown that elimination of misspecified cells due to reduced Dpp signaling is achieved by the interaction of the co-repressor NAB with the transcriptional repressor Brk, which in turn induces Jun N-terminal kinase-dependent apoptosis. Here, we performed a systematic in vivo loss- and gain-of-function analysis to study NAB-induced death and CP. Our findings indicate that the NAB primary signal activates JNK, which in turn transmits two independent signals. One triggers apoptosis through the pro-apoptotic proteins Reaper and Hid, which in turn promote activation of caspases by the apoptosome components Ark and Dronc. The other signal induces CP in a manner that is independent of the death signal, Dronc, or Dp53. Once induced, the apoptotic pathway further activates a CP response. Our data suggest that JNK is the candidate factor that differentiates between apoptosis that involves CP and apoptosis that does not.

Interference in xbp1 gene expression induces defective cell differentiation and sensory organ development in Drosophila

The X-box binding protein 1 (Xbp1) is a transcription factor that is important in the unfolded protein response to protect cells from endoplasmic reticulum (ER) stress and is implicated in several other biological processes, including liver growth and B lymphocyte differentiation. Although the cellular function of Xbp1 has been well studied, its developmental role remains to be elucidated. In the present study, we investigated the developmental role of Drosophila xbp1 using an RNAi-mediated gene silencing system. When xbp1 gene expression was specifically interfered with in developing tissues, such as wing and eye during larval growth, the tissues were strongly deformed. Cell death was not observed in these tissues, and this phenotype was not rescued by the overexpression of Drosophila inhibitor of apoptosis protein1 (DIAP1), a caspase inhibitor, indicating that these developmental defects are not associated with apoptosis. Furthermore, changes were not observed in cell cycle progression and Jun-N-terminal kinase activation in tissues whose xbp1 gene expression was interfered with. Rather, the xbp1 gene silencing resulted in defective cell differentiation and supernumerary of sensory organ precursors. These results suggest the inability of xbp1 gene silencing to induce strong ER stress and the importance of Drosophila Xbp1 in cell fate determination and sensory organ development.

Improved Activities of CREB Binding Protein, Heterogeneous Nuclear Ribonucleoproteins and Proteasome Following Downregulation of Noncoding hsrω Transcripts Help Suppress Poly(Q) Pathogenesis in Fly Models

Following earlier reports on modulation of poly(Q) toxicity in Drosophila by the developmentally active and stress-inducible noncoding hsromega gene, we investigated possible mediators of this modulation. RNAi-mediated downregulation of the large nuclear hsromega-n transcript, which organizes the nucleoplasmic omega speckles, suppressed the enhancement of poly(Q) toxicity brought about by reduced availability of the heterogeneous nuclear ribonucleoprotein (hnRNP) Hrb87F and of the transcriptional regulator, cAMP response element binding (CREB) binding protein (CBP). Levels of CBP RNA and protein were reciprocally affected by hsromega transcript levels in eye disc cells. Our data suggest that CBP and hnRNPs like Hrb57A and Hrb87F physically interact with each other. In addition, downregulation of hsromega transcripts partially rescued eye damage following compromised proteasome activity, while overexpression of hsromega and/or poly(Q) proteins disrupted the proteasomal activity. Rescue of poly(Q) toxicity by hsromega-RNAi required normal proteasomal function. We suggest that hsromega-RNAi suppresses poly(Q) toxicity by elevating cellular levels of CBP, by enhancing proteasome-mediated clearance of the pathogenic poly(Q) aggregates, and by inhibiting induced apoptosis. The direct and indirect interactions of the hsromega transcripts with a variety of regulatory proteins like hnRNPs, CBP, proteasome, Drosophila inhibitor of apoptosis protein 1 (DIAP1), etc., reinforce the view that the noncoding hsromega RNA functions as a "hub" in cellular networks to maintain homeostasis by coordinating the functional availability of crucial cellular regulatory proteins.

The Developmentally Active and Stress-Inducible Noncoding hsrω Gene Is a Novel Regulator of Apoptosis in Drosophila

The large nucleus limited noncoding hsromega-n RNA of Drosophila melanogaster is known to associate with a variety of heterogeneous nuclear RNA-binding proteins (hnRNPs) and certain other RNA-binding proteins to assemble the nucleoplasmic omega speckles. In this article, we show that RNAi-mediated depletion of this noncoding RNA dominantly suppresses apoptosis, in eye and other imaginal discs, triggered by induced expression of Rpr, Grim, or caspases (initiator as well as effector), all of which are key regulators/effectors of the canonical caspase-mediated cell death pathway. We also show, for the first time, a genetic interaction between the noncoding hsromega transcripts and the c-Jun N-terminal kinase (JNK) signaling pathway since downregulation of hsromega transcripts suppressed JNK activation. In addition, hsromega-RNAi also augmented the levels of Drosophila Inhibitor of Apoptosis Protein 1 (DIAP1) when apoptosis was activated. Suppression of induced cell death following depletion of hsromega transcripts was abrogated when the DIAP1-RNAi transgene was coexpressed. Our results suggest that the hsromega transcripts regulate cellular levels of DIAP1 via the hnRNP Hrb57A, which physically interacts with DIAP1, and any alteration in levels of the hsromega transcripts in eye disc cells enhances association between these two proteins. Our studies thus reveal a novel regulatory role of the hsromega noncoding RNA on the apoptotic cell death cascade through multiple paths. These observations add to the diversity of regulatory functions that the large noncoding RNAs carry out in the cells' life.

DXNP/DATRX increases apoptosis via the JNK and dFOXO pathway in Drosophila neurons

Mutation of the XNP/ATRX gene, which encodes an SNF2 family ATPase/helicase protein, leads to ATR-X syndrome and several other X-linked mental retardation syndromes. Although XNP/ATRX is a chromatin remodeler, the molecular mechanism by which mental retardation occurs in patients with ATR-X has yet to be determined. To better understand the role of XNP/ATRX in neuronal development, we expressed Drosophila XNP ( dXNP/DATRX) ectopically in Drosophila neurons. Neuronal expression of dXNP/DATRX resulted in various developmental defects and induced strong apoptosis. These defects and apoptosis were suppressed by Drosophila inhibitor of apoptosis protein 1. Expression of dXNP/DATRX also increased JNK activity and the levels of reaper and hid transcripts, which are pro-apoptotic factors that activate caspase. Furthermore, dXNP/DATRX-induced rough eye phenotype and apoptosis were suppressed by dFOXO deficiency. These results suggest that dXNP/DATRX is involved in caspase-dependent apoptosis in Drosophila neurons via regulation of the JNK and dFOXO pathway.

High levels of dRYBP induce apoptosis in Drosophila imaginal cells through the activation of reaper and the requirement of trithorax, dredd and dFADD

Drosophila RYBP (dRYBP; Ring1 and YY1 Binding Protein) is a Polycomb and trithorax interacting protein, whose homologous RYBP/DEDAF mammalian counterparts exhibit tumor cell-specific killing activity. Here we show that although endogenous dRYBP is not involved in developmental apoptosis, high levels of exogenous dRYBP induce apoptosis in all the imaginal discs of the fly, indicating that dRYBP apoptotic activity is not specific to tumor cells. We also show that dRYBP-induced apoptosis is inhibited by high levels of either p35 or DIAP1 (Drosophila Inhibitor of Apoptosis Protein 1), and requires the function of the pro-apoptotic REAPER, HID and GRIM proteins, the apical caspase DREDD, the adaptor dFADD protein as well as TRITHORAX (TRX), an epigenetic transcriptional regulator. Furthermore, we demonstrate that overexpression of TRX also induces apoptosis in the imaginal discs. Finally, we show that the expression of reaper-lacZ is upregulated both upon dRYBP-induced apoptosis and upon TRX-induced apoptosis in imaginal discs and that the reaper gene is a direct target of dRYBP in Drosophila embryos. Our results indicate that dRYBP triggers in a receptor-mediated apoptotic pathway that also includes TRX-dependent epigenetic regulation of gene expression.

Regulation of the Drosophila apoptosome through feedback inhibition

Apoptosis is induced by caspases, which are members of the cysteine protease family 1. Caspases are synthesized as inactive zymogens and initiator caspases first gain activity by associating with an oligomeric complex of their adaptor proteins, such as the apoptosome 2,3. Activated initiator caspases subsequently cleave and activate effector caspases. While such a proteolytic cascade would predict that a small number of active caspases could irreversibly amplify caspase activity and trigger apoptosis, many cells can maintain moderate levels of caspase activity to perform non-apoptotic roles in cellular differentiation, shape change and migration 4. Here we show that the Drosophila apoptosome engages in a feedback inhibitory loop, thereby moderating its activation level in vivo. Specifically, the adaptor protein Apaf-1 lowers the level of its associated initiator caspase, Dronc, without triggering apoptosis. Conversely, Dronc lowers Apaf-1 protein levels. This mutual suppression depends upon Dronc’s catalytic site and a caspase cleavage site within Apaf-1. Moreover, the Drosophila Inhibitor of Apoptosis Protein 1 (Diap1) is required for this process. We speculate that this feedback inhibition allows cells to regulate the degree of caspase activation for apoptotic and non-apoptotic purposes.

Evolution of mitochondrial cell death pathway: Proapoptotic role of HtrA2/Omi in Drosophila

Despite the essential role of mitochondria in a variety of mammalian cell death processes, the involvement of mitochondrial pathway in Drosophila cell death has remained unclear. To address this, we cloned and characterized DmHtrA2, a Drosophila homolog of a mitochondrial serine protease HtrA2/Omi. We show that DmHtrA2 normally resides in mitochondria and is up-regulated by UV-irradiation. Upon receipt of apoptotic stimuli, DmHtrA2 is translocated to extramitochondrial compartment; however, unlike its mammalian counterpart, the extramitochondrial DmHtrA2 does not diffuse throughout the cytosol but stays near the mitochondria. RNAi-mediated knock-down of DmHtrA2 in larvae or adult flies results in a resistance to stress stimuli. DmHtrA2 specifically cleaves Drosophila inhibitor-of-apoptosis protein 1 (DIAP1), a cellular caspase inhibitor, and induces cell death both in vitro and in vivo as potent as other fly cell death proteins. Our observations suggest that DmHtrA2 promotes cell death through a cleavage of DIAP1 in the vicinity of mitochondria, which may represent a prototype of mitochondrial cell death pathway in evolution.

Identification of E2/E3 Ubiquitinating Enzymes and Caspase Activity Regulating Drosophila Sensory Neuron Dendrite Pruning

Ubiquitin-proteasome system (UPS) is a multistep protein degradation machinery implicated in many diseases. In the nervous system, UPS regulates remodeling and degradation of neuronal processes and is linked to Wallerian axonal degeneration, though the ubiquitin ligases that confer substrate specificity remain unknown. Having shown previously that class IV dendritic arborization (C4da) sensory neurons in Drosophila undergo UPS-mediated dendritic pruning during metamorphosis, we conducted an E2/E3 ubiquitinating enzyme mutant screen, revealing that mutation in ubcD1, an E2 ubiquitin-conjugating enzyme, resulted in retention of C4da neuron dendrites during metamorphosis. Further, we found that UPS activation likely leads to UbcD1-mediated degradation of DIAP1, a caspase-antagonizing E3 ligase. This allows for local activation of the Dronc caspase, thereby preserving C4da neurons while severing their dendrites. Thus, in addition to uncovering E2/E3 ubiquitinating enzymes for dendrite pruning, this study provides a mechanistic link between UPS and the apoptotic machinery in regulating neuronal process remodeling.