Cell Death & Differentiation

Derry, W B; Bierings, R; van Iersel, M; Satkunendran, T; Reinke, V; Rothman, J H

doi: 10.1038/sj.cdd.4402075pmid: 17186023

Caenorhabditis elegans CEP-1 activates germline apoptosis in response to genotoxic stress, similar to its mammalian counterpart, tumor suppressor p53. In mammals, there are three p53 family members (p53, p63, and p73) that activate and repress many distinct and overlapping sets of genes, revealing a complex transcriptional regulatory network. Because CEP-1 is the sole p53 family member in C. elegans, analysis of this network is greatly simplified in this organism. We found that CEP-1 functions during normal development in the absence of stress to repress many (331) genes and activate only a few (28) genes. In response to genotoxic stress, 1394 genes are activated and 942 are repressed, many of which contain p53-binding sites. Comparison of the CEP-1 transcriptional network with transcriptional targets of the human p53 family reveals considerable overlap between CEP-1-regulated genes and homologues regulated by human p63 and p53, suggesting a composite p53/p63 action for CEP-1. We found that phg-1, the C. elegans Gas1 (growth arrest-specific 1) homologue, is activated by CEP-1 and is a negative regulator of cell proliferation in the germline in response to genotoxic stress. Further, we find that CEP-1 and PHG-1 mediate the decreased developmental rate and embryonic viability of mutations in the clk-2/TEL2 gene, which regulates lifespan and checkpoint responses.

Hanoux, V; Pairault, C; Bakalska, M; Habert, R; Livera, G

doi: 10.1038/sj.cdd.4402052pmid: 17082817

In mammals, the pool of primordial follicles at birth is determinant for female fertility. Exposure to IR during oogonia proliferation and the diplotene stages of ovarian development induced the virtual disappearance of primordial follicles in the postnatal ovary, while half the follicular reserve remained present after irradiation during the zygotene/pachytene stages. This sensitivity difference was correlated with the level of caspase-2 expression evaluated by immunohistochemistry. At the diplotene stage, Western blot and caspase activity analysis revealed that caspase-2 was activated 2 h after irradiation and a significant increase in the number of oocytes expressing cleaved caspase-9 and -3 occurred 6 h after treatment. Inhibition of caspase-2 activity prevented the cleavage of caspase-9 and partially prevented the loss of oocytes in response to irradiation. Taken together, our results show that caspase-2-dependent activation of the mitochondrial apoptotic pathway is one of the mechanisms involved in the genotoxic stress-induced depletion of the primordial follicle pool.

Olichon, A; ElAchouri, G; Baricault, L; Delettre, C; Belenguer, P; Lenaers, G

doi: 10.1038/sj.cdd.4402048pmid: 17024226

In most eucaryote cells, release of apoptotic proteins from mitochondria involves fission of the mitochondrial network and drastic remodelling of the cristae structures. The intramitochondrial dynamin OPA1, as a potential central actor of these processes, exists as eight isoforms resulting from the alternate splicing combinations of exons (Ex) 4, 4b and 5b, which functions remain undetermined. Here, we show that Ex4 that is conserved throughout evolution confers functions to OPA1 involved in the maintenance of the ΔΨm and in the fusion of the mitochondrial network. Conversely, Ex4b and Ex5b, which are vertebrate specific, define a function involved in cytochrome c release, an apoptotic process also restricted to vertebrates. The drastic changes of OPA1 variant abundance in different organs suggest that nuclear splicing can control mitochondrial dynamic fate and susceptibility to apoptosis and pathologies.

Comes, F; Matrone, A; Lastella, P; Nico, B; Susca, F C; Bagnulo, R; Ingravallo, G; Modica, S; Lo Sasso, G; Moschetta, A; Guanti, G; Simone, C

doi: 10.1038/sj.cdd.4402076pmid:

Zhang, D; Armstrong, J S

doi: 10.1038/sj.cdd.4402072pmid: 17170750

Endoplasmic reticulum (ER) stress induces apoptosis by mechanisms that are not fully clear. Here we show that ER stress induced by the Ca2+-ATPase inhibitor thapsigargin (THG) activates cytochrome c-dependent apoptosis through cooperation between Bax and the mitochondrial permeability transition (MPT) in human leukemic CEM cells. Pharmacological inhibition of the MPT as well as small interfering RNA (siRNA) knockdown of the MPT core component cyclophilin D blocked cytochrome c release and caspase-dependent apoptosis but did not prevent Bax activation, translocation or N-terminal exposure in mitochondria. siRNA knockdown of Bax also blocked THG-mediated cytochrome c release and apoptosis, but did not prevent MPT activation and resulted in caspase-independent cell death. Our results show that ER-stress-induced cell death involves a caspase and Bax-dependent pathway as well as a caspase-independent MPT-directed pathway.

Miyata, K; Yasukawa, T; Fukuda, M; Takeuchi, T; Yamazaki, K; Sakumi, K; Tamamori-Adachi, M; Ohnishi, Y; Ohtsuki, Y; Nakabeppu, Y; Kitajima, S; Onishi, S; Aso, T

doi: 10.1038/sj.cdd.4402067

Sexton, K B; Kato, D; Berger, A B; Fonovic, M; Verhelst, S H L; Bogyo, M

doi: 10.1038/sj.cdd.4402074pmid: 17170749

Activity-Based Probes (ABPs) are small molecules that form stable covalent bonds with active enzymes thereby allowing detection and quantification of their activities in complex proteomes. A number of ABPs that target proteolytic enzymes have been designed based on well-characterized mechanism-based inhibitors. We describe here the evaluation of a novel series of ABPs based on the aza-aspartate inhibitory scaffold. Previous in vitro kinetic studies showed that this scaffold has a high degree of selectivity for the caspases, clan CD cysteine proteases activated during apoptotic cell death. Aza-aspartate ABPs containing either an epoxide or Michael acceptor reactive group were potent labels of executioner caspases in apoptotic cell extracts. However they were also effective labels of the clan CD protease legumain and showed unexpected crossreactivity with the clan CA protease cathepsin B. Interestingly, related aza peptides containing an acyloxymethyl ketone reactive group were relatively weak but highly selective labels of caspases. Thus azapeptide electrophiles are valuable new ABPs for both detection of a broad range of cysteine protease activities and for selective targeting of caspases. This study also highlights the importance of confirming the specificity of covalent protease inhibitors in crude proteomes using reagents such as the ABPs described here.

Franz, S; Herrmann, K; Führnrohr, B; Sheriff, A; Frey, B; Gaipl, U S; Voll, R E; Kalden, J R; Jäck, H-M; Herrmann, M

doi: 10.1038/sj.cdd.4402066pmid: 17170754

Apoptosis and phagocytosis of apoptotic cells are crucial processes. At best the phagocytic machinery detects and swallows all apoptotic cells in a way that progression to secondary necrosis is avoided. Otherwise, inflammation and autoimmune diseases may occur. Most apoptotic cells are phagocytosed instantaneously in a silent fashion; however, some dying cells escape their clearance. If the cells are not cleared early, they lose membranes due to extensive shedding of membrane surrounded vesicles (blebbing) and shrink. It is unclear how apoptotic cells compensate their massive loss of plasma membrane. Here, we demonstrate that endoplasmic reticulum- (ER) resident proteins (calnexin, the KDEL receptor and a dysfunctional immunoglobulin heavy chain) were exposed at the surfaces of shrunken late apoptotic cells. Additionally, these cells showed an increased binding of lectins, which recognize sugar structures predominantly found as moieties of incompletely processed proteins in ER and Golgi. In addition the ER resident lipophilic ER-Tracker™ Blue-White DPX, and internal GM1 were observed to translocate to the cell surfaces during late apoptosis. We conclude that during blebbing of apoptotic cells the surface membrane loss is substituted by immature membranes from internal stores. This mechanism explains the simultaneous appearance of preformed recognition structures for several adaptor proteins known to be involved in clearance of dead cells.

Levy, D; Adamovich, Y; Reuven, N; Shaul, Y

doi: 10.1038/sj.cdd.4402063pmid: 17110958

Upon DNA damage signaling, p73, a member of the p53 tumor suppressor family, accumulates to support transcription of downstream apoptotic genes. p73 interacts with Yes-associated protein 1 (Yap1) through its PPPY motif, and increases p73 transactivation of apoptotic genes. The ubiquitin E3 ligase Itch, like Yap1, interacts with p73. Given the fact that both Itch and Yap1 bind p73 via the PPPY motif, we hypothesized that Yap may also function to stabilize p73 by displacing Itch binding to p73. We show that the interaction of Yap1 and p73 was necessary for p73 stabilization. Yap1 competed with Itch for binding to p73, and prevented Itch-mediated ubiquitination of p73. Treatment of cells with cisplatin leads to an increase in p73 accumulation and induction of apoptosis, but both were dramatically reduced in the presence of Yap1 siRNA. Altogether, our findings attribute a central role to Yap1 in regulating p73 accumulation and function under DNA damage signaling.

Foveau, B; Leroy, C; Ancot, F; Deheuninck, J; Ji, Z; Fafeur, V; Tulasne, D

doi: 10.1038/sj.cdd.4402080pmid: 17186028

Activation of the MET tyrosine kinase receptor by hepatocyte growth factor/scatter factor is classically associated with cell survival. Nonetheless, stress stimuli can lead to a caspase-dependent cleavage of MET within its juxtamembrane region, which generate a proapoptotic 40 kDa fragment (p40 MET). We report here that p40 MET is in fact generated through an additional caspase cleavage of MET within its extreme C-terminal region, which removes only few amino acids. We evidenced a hierarchical organization of these cleavages, with the C-terminal cleavage favoring the juxtamembrane one. As a functional consequence, the removal of the last amino acids of p40 MET increases its apoptotic capacity. Finally, cells expressing a MET receptor mutated at the C-terminal caspase site are unable to generate p40 MET and are resistant to apoptosis, indicating that generation of p40 MET amplifies apoptosis. These results revealed a two-step caspase cleavage of MET resulting in the reshaping of this survival receptor to a proapoptotic factor.