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Insects have made major contributions to understanding the regulation of cell death, dating back to the pioneering work of Lockshin and Williams on death of muscle cells during postembryonic development of Manduca . A physically smaller cousin of moths, the fruit fly Drosophila melanogaster, offers unique advantages for studying the regulation of cell death in response to different apoptotic stimuli in situ. Different signaling pathways converge in Drosophila to activate a common death program through transcriptional activation of reaper, hid and grim. Reaper-family proteins induce apoptosis by binding to and antagonizing inhibitor of apoptosis proteins (IAPs), which in turn inhibit caspases. This switch from life to death relies extensively on targeted degradation of cell death proteins by the ubiquitin–proteasome pathway. Drosophila IAP-1 (Diap1) functions as an E3-ubiquitin ligase to protect cells from unwanted death by promoting the degradation of the initiator caspase Dronc. However, in response to apoptotic signals, Reaper-family proteins are produced, which promote the auto-ubiquitination and degradation of Diap1, thereby removing the ‘brakes on death’ in cells that are doomed to die. More recently, several other ubiquitin pathway proteins were found to play important roles for caspase regulation, indicating that the control of cell survival and death relies extensively on targeted degradation by the ubiquitin–proteasome pathway.

Programmed cell death is an important process during development that serves to remove superfluous cells and tissues, such as larval organs during metamorphosis, supernumerary cells during nervous system development, muscle patterning and cardiac morphogenesis. Different kinds of cell death have been observed and were originally classified based on distinct morphological features: (1) type I programmed cell death (PCD) or apoptosis is recognized by cell rounding, DNA fragmentation, externalization of phosphatidyl serine, caspase activation and the absence of inflammatory reaction, (2) type II PCD or autophagy is characterized by the presence of large vacuoles and the fact that cells can recover until very late in the process and (3) necrosis is associated with an uncontrolled release of the intracellular content after cell swelling and rupture of the membrane, which commonly induces an inflammatory response. In this review, we will focus exclusively on developmental cell death by apoptosis and its role in tissue remodeling.

Cell death is essential for a plethora of physiological processes, and its deregulation characterizes numerous human diseases. Thus, the in-depth investigation of cell death and its mechanisms constitutes a formidable challenge for fundamental and applied biomedical research, and has tremendous implications for the development of novel therapeutic strategies. It is, therefore, of utmost importance to standardize the experimental procedures that identify dying and dead cells in cell cultures and/or in tissues, from model organisms and/or humans, in healthy and/or pathological scenarios. Thus far, dozens of methods have been proposed to quantify cell death-related parameters. However, no guidelines exist regarding their use and interpretation, and nobody has thoroughly annotated the experimental settings for which each of these techniques is most appropriate. Here, we provide a nonexhaustive comparison of methods to detect cell death with apoptotic or nonapoptotic morphologies, their advantages and pitfalls. These guidelines are intended for investigators who study cell death, as well as for reviewers who need to constructively critique scientific reports that deal with cellular demise. Given the difficulties in determining the exact number of cells that have passed the point-of-no-return of the signaling cascades leading to cell death, we emphasize the importance of performing multiple, methodologically unrelated assays to quantify dying and dead cells.

The encapsulation of genetic polymers inside lipid bilayer compartments (vesicles) is a vital step in the emergence of cell-based life. However, even though acidic conditions promote many reactions required for generating prebiotic building blocks, prebiotically relevant lipids tend to form denser aggregates at acidic pHs rather than prebiotically useful vesicles that exhibit sufficient solute encapsulation. Here, we describe how dehydration/rehydration (DR) events, a prebiotically relevant physicochemical process known to promote polymerization reactions, can remodel dense lipid aggregates into thin-walled vesicles capable of RNA encapsulation even at acidic pHs. Furthermore, DR events appear to favor the encapsulation of RNA within thin-walled vesicles over more lipid-rich vesicles, thus conferring such vesicles a selective advantage.

Programmed cell death (PCD) in the Drosophila ovary occurs either during mid-oogenesis, resulting in degeneration of the entire egg chamber or during late oogenesis, to facilitate the development of the oocyte. PCD during oogenesis is regulated by mechanisms different from those that control cell death in other Drosophila tissues. We have analyzed the role of caspases in PCD of the female germline by examining caspase mutants and overexpressing caspase inhibitors. Imprecise P- element excision was used to generate mutants of the initiator caspase strica. While null mutants of strica or another initiator caspase, dronc, display no ovary phenotype, we find that strica exhibits redundancy with dronc , during both mid- and late oogenesis. Ovaries of double mutants contain defective mid-stage egg chambers similar to those reported previously in dcp-1 mutants, and mature egg chambers with persisting nurse cell nuclei. In addition, the effector caspases drice and dcp-1 also display redundant functions during late oogenesis, resulting in persisting nurse cell nuclei. These findings indicate that caspases are required for nurse cell death during mid-oogenesis, and participate in developmental nurse cell death during late oogenesis. This reveals a novel pathway of cell death in the ovary that utilizes strica , dronc , dcp-1 and drice , and importantly illustrates strong redundancy among the caspases.

The mammalian p53 family consists of p53, p63 and p73. Whereas p53 accounts for tumor suppression through cell-cycle arrest and apoptosis, the functions of p63 and p73 are more diverse and also include control of cell differentiation. The Drosophila genome contains only one p53 homolog, Dp53 . Previous work has established that Drosophila p53 (Dp53) induces apoptosis, but not cell-cycle arrest. In this study, using the developing eye as a model, we show that Dp53 -induced apoptosis is primarily dependent on the pro-apoptotic gene, head involution defective ( hid ), but not reaper ( rpr ), and occurs through the canonical apoptosis pathway. Importantly, similar to p63 and p73, expression of Dp53 also inhibits cellular differentiation of photoreceptor neurons and cone cells in the eye independently of its apoptotic function. Intriguingly, expression of the human cell-cycle inhibitor p21 or its Drosophila homolog dacapo ( dap ) can suppress both Dp53-induced cell death and differentiation defects in Drosophila eyes. These findings provide new insights into the pathways activated by Dp53 and reveal that Dp53 incorporates functions of multiple p53 family members.

In Drosophila, the chromosomal region 75C1-2 contains at least three genes, reaper (rpr), head involution defective (hid), and grim, that have important functions in the activation of programmed cell death. To better understand how cells are killed by these genes, we have utilized a well defined set of embryonic central nervous system midline cells that normally exhibit a specific pattern of glial cell death. In this study we show that both rpr and hid are expressed in dying midline cells and that the normal pattern of midline cell death requires the function of multiple genes in the 75C1-2 interval. We also utilized the P[UAS]/P[Gal4] system to target expression of rpr and hid to midline cells. Targeted expression of rpr or hid alone was not sufficient to induce ectopic midline cell death. However, expression of both rpr and hid together rapidly induced ectopic midline cell death that resulted in axon scaffold defects characteristic of mutants with abnormal midline cell development. Midline-targeted expression of the baculovirus p35 protein, a caspase inhibitor, blocked both normal and ectopic rpr- and hid-induced cell death. Taken together, our results suggest that rpr and hid are expressed together and cooperate to induce programmed cell death during development of the central nervous system midline.

Apoptosis is a morphologically distinct form of programmed cell death that plays a major role during development, homeostasis, and in many diseases including cancer, acquired immunodeficiency syndrome, and neurodegenerative disorders. Apoptosis occurs through the activation of a cell-intrinsic suicide program. The basic machinery to carry out apoptosis appears to be present in essentially all mammalian cells at all times, but the activation of the suicide program is regulated by many different signals that originate from both the intracellular and the extracellular milieu. Genetic studies in the nematode Caenorhabditis elegans and in the fruit fly Drosophila melanogaster have led to the isolation of genes that are specifically required for the induction of programmed cell death. At least some components of the apoptotic program have been conserved among worms, insects, and vertebrates.

Three genes--reaper, grim, and his--are crucial to the regulation of programmed cell death in Drosophila melanogaster. Mutations involving all three genes virtually abolish apoptosis during development, and homozygous hid mutants die as embryos with extensive defects in apoptosis. Although Hid is central to apoptosis in Drosophila, it has no mammalian homologue identified to date. We present evidence that expression of Drosophila Hid in mammalian cells induces apoptosis. This activity is subject to regulation by inhibitors of mammalian cell death. We show that the N terminus of Hid, which is a region of homology with Reaper and Grim, is essential for Hid's function in mammalian cells. We demonstrate that Hid is localized to the mitochondria via a hydrophobic region at its C terminus and functionally interacts with BclXL. This study shows that the function of Hid as a death inducer in Drosophila is conserved in mammalian cells and argues for the existence of a mammalian homologue of this critical regulator of apoptosis.