Bcl-xS and Bax induce different apoptotic pathways in PC12 cells

Lindenboim, Liora; Yuan, Junying; Stein, Reuven

doi:10.1038/sj.onc.1203495

Cited by 69 publications

(54 citation statements)

References 66 publications

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“…The expression vectors pEGFP, pcDNA3, pCMV-SEAP, and pRSV-H-Ras(S17N) (DN-Ras) have been described elsewhere. 35,36 The MSCV-NF1GRD vector and its empty puromycin resistance gene (Pac) vector were obtained from Dr. D Wade Clapp (Indiana University School of Medicine).…”

Section: Methodsmentioning

confidence: 99%

“…SEAP activity was assayed as described elsewhere. 35 Briefly, 200 ml from the culture medium of the transfected cells were collected and spun for 2 min at 10 000 Â g. The supernatant was incubated at 651C for 10 min, and aliquots (30 ml) from each treatment were then incubated with 200 ml of SEAP buffer (1 M diethanolamine, 0.5 mM MgCl 2 , 10 mM L-homo-arginine, and 5 mg/ml p-nitrophenyl phosphate; at 371C for about 4 h. Assays were performed in quadruplicate for each treatment. The plates were read on a microELISA reader at a wavelength of 405 nm.…”

Section: Assessment Of Cell Survivalmentioning

confidence: 99%

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The tumor suppressor neurofibromin confers sensitivity to apoptosis by Ras-dependent and Ras-independent pathways

et al. 2006

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Neurofibromatosis type 1 (NF1) is characterized by a high incidence of benign and malignant tumors attributed to loss of function of Nf1, which encodes neurofibromin, a tumor suppressor with Ras-GAP activity. Neurofibromin deficiency typically causes chronic activation of Ras, considered the major contributor to manifestation of NF1. Resistance to radio-and chemotherapy are typical of NF1-associated tumors, but the underlying mechanism is unknown. Here, we investigated interrelationships between neurofibromin expression, Ras activity, and sensitivity to apoptosis. Neurofibromin-deficient mouse embryonic fibroblasts (MEFs) and human NF1 tumor cells were more resistant than neurofibromin-expressing cells to apoptosis. Moreover, Nf1 À/À , Nf1 þ /À , and Nf1 þ / þ MEFs exhibited gene-dosage-related resistance to apoptosis. Resistance of the Nf1-deficient cells was mediated by two survival pathways: a Ras-dependent pathway, and a Ras-independent pathway promoted by the lack of an NF1-GRD-independent proapoptotic action of neurofibromin. Therefore, besides its Ras-dependent growth inhibition, neurofibromin can exert tumor suppression via a proapoptotic effect.

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Section: Methodsmentioning

confidence: 99%

Section: Assessment Of Cell Survivalmentioning

confidence: 99%

The tumor suppressor neurofibromin confers sensitivity to apoptosis by Ras-dependent and Ras-independent pathways

et al. 2006

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“…Caspase-independent apoptosis has been described in several other models of apoptosis including cell death induced by vitamin D compounds 44 nitric oxide, 9 depletion of heat shock protein 70, 45 Bax, 46 Bak, 33 and the c-Myc interacting adaptor protein Bin 1. 47 Furthermore, this laboratory has previously described a retinal cell apoptosis pathway in vitro which does not involve caspase activity and retains key features of apoptotic cell death including DNA strand nicking, phosphatidylserine externalisation and cell shrinkage.…”

Section: Discussionmentioning

confidence: 99%

Caspase-independent photoreceptor apoptosis in vivo and differential expression of apoptotic protease activating factor-1 and caspase-3 during retinal development

2002

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Apoptosis is the mode of photoreceptor cell death in many retinal dystrophies. Exposure of Balb/c mice to excessive levels of light induces photoreceptor apoptosis and represents an animal model for the study of retinal degenerations. Caspases have emerged as central regulators of apoptosis, executing this tightly controlled death pathway in many cells. Previously we have reported that light-induced photoreceptor apoptosis occurs independently of one the key executioners of apoptosis, caspase-3. This present study extends these results reporting on the lack of activation of other caspases in this model including caspases-8, -9, -7, and -1. Furthermore, photoreceptor apoptosis cannot be inhibited with the broad range caspase inhibitor zVAD-fmk indicating that lightinduced retinal degeneration is caspase-independent. We demonstrate that cytochrome c does not translocate from mitochondria to the cytosol during photoreceptor apoptosis. We also show that during retinal development apoptotic protease activating factor (Apaf-1) protein levels are markedly decreased and this is associated with the inability to activate the mitochondrial caspase cascade in the mature retina. In addition, there is also a significant reduction in expression of caspases-3 and -9 during retinal maturation and these levels do not increase following light exposure. Finally, we show that the calcium-dependent proteases calpains are active during light-induced retinal degeneration and establish that the calcium channel blocker D-cis-diltiazem completely inhibits photoreceptor apoptosis.

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“…Proapoptotic Bcl-2 family members such as Bax and Bcl-X S have been shown to be activated and translocated from the cytosol into mitochondria after apoptotic stimulation. 24,38 Since human a-crystallins were able to interact with Bax and Bcl-X S in vitro and in vivo, we predicted that the translocation of Bax and Bcl-X S could be affected in the presence of HaA and HaB. To test this possibility, the pEGFP-HLE, pEGFP-HaA-HLE and pEGFPHaB-HLE cells were treated with either 0.01% DMSO or 100 nM staurosporine for 3 h and the mitochondrial-associated (M) and soluble (S) proteins were isolated from the three types of cells as described in Materials and Methods.…”

Section: Haa-and Hab-crystallins Can Interact With Bcl-x S and Bax Inmentioning

confidence: 99%

“…Its proapoptotic ability can be inhibited by Bcl-2 and Bcl-X L . 38 Another important group of apoptosis regulators are heatshock proteins that include alpha-crystallins (a-crystallins), [39][40][41][42][43][44][45][46] Hsp27, [39][40][47][48][49][50][51][52] Hsp70, [53][54][55][56][57] Hsp90 58 and Hsp60. [59][60] While Hsp60 appears to enhance apoptosis by promoting maturation of procaspase-3, 59,60 the majority of these factors seems to protect cells from induced apoptosis through different mechanisms.…”

Section: Introductionmentioning

confidence: 99%

Human αA- and αB-crystallins bind to Bax and Bcl-XS to sequester their translocation during staurosporine-induced apoptosis

et al. 2004

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aA-and aB-crystallins are distinct antiapoptotic regulators. Regarding the antiapoptotic mechanisms, we have recently demonstrated that aB-crystallin interacts with the procaspase-3 and partially processed procaspase-3 to repress caspase-3 activation. Here, we demonstrate that human aA-and aBcrystallins prevent staurosporine-induced apoptosis through interactions with members of the Bcl-2 family. Using GST pulldown assays and coimmunoprecipitations, we demonstrated that a-crystallins bind to Bax and Bcl-X S both in vitro and in vivo. Human aA-and aB-crystallins display similar affinity to both proapoptotic regulators, and so are true with their antiapoptotic ability tested in human lens epithelial cells, human retina pigment epithelial cells (ARPE-19) and rat embryonic myocardium cells (H9c2) under treatment of staurosporine, etoposide or sorbitol. Two prominent mutants, R116C in aA-crystallin and R120G, in aB-crystallin display much weaker affinity to Bax and Bcl-X S . Through the interaction, a-crystallins prevent the translocation of Bax and Bcl-X S from cytosol into mitochondria during staurosporine-induced apoptosis. As a result, a-crystallins preserve the integrity of mitochondria, restrict release of cytochrome c, repress activation of caspase-3 and block degradation of PARP. Thus, our results demonstrate a novel antiapoptotic mechanism for a-crystallins. Keywords: aA-crystallin; R116C; aB-crystallin; R120G; Bax; Bcl-X S Abbreviations: a-crystallin, alpha-crystallin; GFP, green fluorescence protein; HaA, human aA-crystallin; HaB, human aB-crystallin; GFP-HaA, green fluorescence and human aAcrystallin fusion protein; GFP-HaB, green fluorescence and human aB-crystallin fusion protein; MEM, Eagle's minimal essential medium; PAGE, polyacrylamide gel electrophoresis; PBS, phosphate-buffered saline; PMSF, phenylmethylsufonyl fluoride; HLE, human lens epithelial cells; SDS, sodium dodecylsulfate; TBS, tris-buffered saline; TBS-T, tris-buffered saline with tween-20.

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Bcl-xS and Bax induce different apoptotic pathways in PC12 cells

Cited by 69 publications

References 66 publications

The tumor suppressor neurofibromin confers sensitivity to apoptosis by Ras-dependent and Ras-independent pathways

The tumor suppressor neurofibromin confers sensitivity to apoptosis by Ras-dependent and Ras-independent pathways

Caspase-independent photoreceptor apoptosis in vivo and differential expression of apoptotic protease activating factor-1 and caspase-3 during retinal development

Human αA- and αB-crystallins bind to Bax and Bcl-XS to sequester their translocation during staurosporine-induced apoptosis

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