BAD Enables Ceramide to Signal Apoptosis via Ras and Raf-1

Abstract

Prior investigations document that proliferative signaling cascades, under some circumstances, initiate apoptosis, although mechanisms that dictate the final outcome are largely unknown. In COS-7 cells, ceramide signals Raf-1 activation through Ras (Zhang, Y., Yao, B., Delikat, S., Bayoumy, S., Lin, X. H., Basu, S., McGinley, M., Chan-Hui, P. Y., Lichenstein, H., and Kolesnick, R. (1997) Cell 89, 63–72), but not apoptosis. However, expression of small amounts of the pro-apoptotic Bcl-2 family member, BAD, conferred ceramide-induced apoptosis onto COS-7 cells. Ceramide signaled apoptosis in BAD-expressing cells by a pathway involving sequentially kinase suppressor ofRas (KSR)/ceramide-activated protein kinase, Ras, c-Raf-1, and MEK1. Downstream, this pathway linked to BAD dephosphorylation at serine 136 by prolonged inactivation of Akt/PKB. Further, mutation of BAD at serine 136 abrogated ceramide signaling of apoptosis. The present study indicates that when ceramide signals through the Ras/Raf cascade, the availability of a single target, BAD, may dictate an apoptotic outcome. Prior investigations document that proliferative signaling cascades, under some circumstances, initiate apoptosis, although mechanisms that dictate the final outcome are largely unknown. In COS-7 cells, ceramide signals Raf-1 activation through Ras (Zhang, Y., Yao, B., Delikat, S., Bayoumy, S., Lin, X. H., Basu, S., McGinley, M., Chan-Hui, P. Y., Lichenstein, H., and Kolesnick, R. (1997) Cell 89, 63–72), but not apoptosis. However, expression of small amounts of the pro-apoptotic Bcl-2 family member, BAD, conferred ceramide-induced apoptosis onto COS-7 cells. Ceramide signaled apoptosis in BAD-expressing cells by a pathway involving sequentially kinase suppressor ofRas (KSR)/ceramide-activated protein kinase, Ras, c-Raf-1, and MEK1. Downstream, this pathway linked to BAD dephosphorylation at serine 136 by prolonged inactivation of Akt/PKB. Further, mutation of BAD at serine 136 abrogated ceramide signaling of apoptosis. The present study indicates that when ceramide signals through the Ras/Raf cascade, the availability of a single target, BAD, may dictate an apoptotic outcome. tumor necrosis factor kinase suppressor of Ras ceramide-activated protein kinase mitogen-activated protein kinase kinase-inactive KSR phosphatidylinositol polyacrylamide gel electrophoresis protein kinase B. It is generally accepted that a family of evolutionarily conserved cysteine aspartate proteases or caspases serve as effectors of the apoptotic response (1Salvesen G. Dixit V. Cell. 1997; 91: 443-446Abstract Full Text Full Text PDF PubMed Scopus (1936) Google Scholar). The upstream elements that lead to this committed stage are presently the focus of intensive investigation. For cytokine receptors of the tumor necrosis factor (TNF)1 receptor superfamily, a death domain adaptor protein system, activated upon ligand binding, appears to initiate the apoptotic response by linking directly to caspases. For stresses that signal independent of cytokine receptors, the mechanisms of integration into the effector caspase system are less clear. One pathway that appears to be a generic upstream signaling mechanism for induction of apoptosis is the sphingomyelin signaling system (2Pena L.A. Fuks Z. Kolesnick R. Biochem. Pharmacol. 1997; 53: 615-621Crossref PubMed Scopus (220) Google Scholar,3Smyth M.J. Obeid L.M. Hannun Y.A. Adv. Pharmacol. 1997; 41: 133-154Crossref PubMed Scopus (62) Google Scholar). Ceramide is the second messenger of this pathway and is generated by hydrolysis of plasma membrane sphingomyelin through the action of either a neutral or acidic sphingomyelinase, or by de novosynthesis via the enzyme ceramide synthase. The sphingomyelin pathway induces differentiation, proliferation or senescence depending on the cell type. Very often, however, the outcome of signaling through this pathway is apoptosis. Several lines of evidence support the role of ceramide as a signal for the apoptotic response. In a variety of cell types, ceramide generation precedes morphological and biochemical manifestations of apoptosis. Further, specific inhibitors of ceramide synthase, such as the fungal toxin fumonisin B1, antagonize apoptosis mediated by de novoceramide synthesis (4Bose R. Verheij M. Haimovitz-Friedman A. Scotto K. Fuks Z. Kolesnick R. Cell. 1995; 82: 405-414Abstract Full Text PDF PubMed Scopus (784) Google Scholar). Perhaps most convincingly, in systems where the acid sphingomyelinase appears to regulate ceramide signaling of apoptosis such as in peripheral B cells or endothelium, genetic deficiency of acid sphingomyelinase compromises ceramide generation and apoptosis (5Santana P. Pena L.A. Haimovitz-Friedman A. Martin S. Green D. McLoughlin M. Cordon-Cardo C. Schuchman E.H. Fuks Z. Kolesnick R. Cell. 1996; 86: 189-199Abstract Full Text Full Text PDF PubMed Scopus (726) Google Scholar, 6Herr I. Wilhelm D. Bohler T. Angel P. Debatin K.M. EMBO J. 1997; 16: 6200-6208Crossref PubMed Scopus (279) Google Scholar). Recent studies in Saccharomyces cerevisiae revealed that the heat stress response is mediated by synthesis of sphingolipids (7Mandala S.M. Thornton R. Tu Z. Kurtz M.B. Nickels J. Broach J. Menzeleev R. Spiegel S. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 150-155Crossref PubMed Scopus (235) Google Scholar, 8Jenkins G.M. Richards A. Wahl T. Mao C. Obeid L. Hannun Y. J. Biol. Chem. 1997; 272: 32566-32572Abstract Full Text Full Text PDF PubMed Scopus (258) Google Scholar, 9Dickson R.C. Nagiec E.E. Wells G.B. Nagiec M.M. Lester R.L. J. Biol. Chem. 1997; 272: 29620-29625Abstract Full Text Full Text PDF PubMed Scopus (146) Google Scholar). The development of thermotolerance is defective in mutants incapable of synthesizing ceramide, and sphingolipid analogs bypass the defect, restoring growth at elevated temperature. These data indicate that ceramide signaling is evolutionarily older than the caspase/apoptotic death response system, and hence these two pathways must have been linked later in development. Ceramide-induced apoptosis occurs via two independent mechanisms. Ceramide signals, perhaps through the Jun kinase cascade, the transcriptional regulation of gene products, such as Fas ligand or TNFα, that mediate the death response (6Herr I. Wilhelm D. Bohler T. Angel P. Debatin K.M. EMBO J. 1997; 16: 6200-6208Crossref PubMed Scopus (279) Google Scholar, 10Brenner B. Koppenhoefer U. Weinstock C. Linderkamp O. Lang F. Gulbins E. J. Biol. Chem. 1997; 272: 22173-22181Abstract Full Text Full Text PDF PubMed Scopus (297) Google Scholar). Alternately, ceramide induces apoptosis directly through a mechanism inhibitable by anti-apoptotic Bcl-2 family members (11Susin S.A. Zamzami N. Castedo M. Daugas E. Wang H.G. Geley S. Fassy F. Reed J.C. Kroemer G. J. Exp. Med. 1997; 186: 25-37Crossref PubMed Scopus (589) Google Scholar, 12Smyth M.J. Perry D.K. Zhang J. Poirier G.G. Hannun Y.A. Obeid L.M. Biochem. J. 1996; 316: 25-28Crossref PubMed Scopus (198) Google Scholar, 13Farschon D.M. Couture C. Mustelin T. Newmeyer D.D. J. Cell Biol. 1997; 137: 1117-1125Crossref PubMed Scopus (36) Google Scholar). This transcriptionally independent pathway for induction of apoptosis has been observed in cytoplasts (14Castedo M. Hirsch T. Susin S.A. Zamzami N. Marchetti P. Macho A. Kroemer G. J. Immunol. 1996; 157: 512-521PubMed Google Scholar) and recently reconstituted in a cell-free system (15Susin S. Zamzami N. Larochette N. Dallaporta B. Marzo I. Brenner C. Hirsch T. Petit P. Geuskens M. Kroemer G. Exp. Cell Res. 1997; 236: 397-403Crossref PubMed Scopus (75) Google Scholar). The target for ceramide and the signaling system involved in this latter death paradigm are unknown. Recent investigations have begun to elucidate a pathway for induction of apoptosis through the pro-apoptotic Bcl-2 family member BAD (16Wang H.-G. Rapp U.R. Reed J.C. Cell. 1996; 87: 629-638Abstract Full Text Full Text PDF PubMed Scopus (708) Google Scholar, 17Zha J. Harada H. Yang E. Jockel J. Korsmeyer S.J. Cell. 1996; 87: 619-628Abstract Full Text Full Text PDF PubMed Scopus (2253) Google Scholar, 18Datta S.R. H. X. S. H. Y. Cell. 1997; 91: Full Text Full Text PDF PubMed Scopus Google Scholar, L. M. C. R. G. 1997; PubMed Scopus Google Scholar). at the of B cell is in and apoptosis by a variety of A. 1997; Google Scholar, G. Med. 1997; PubMed Scopus Google Scholar, J.C. 1997; PubMed Scopus Google Scholar). mechanisms for the anti-apoptotic action of Bcl-2 family members have been the regulation of the of the for P. H. M. 1997; PubMed Scopus Google for caspase activation E. Green Newmeyer D.D. 1997; PubMed Scopus Google Scholar, P. D. I. S.M. M. Wang X. Cell. 1997; 91: Full Text Full Text PDF PubMed Scopus Google of the G. K. Dixit J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google and and generation Biochem. Sci. 1996; Full Text PDF PubMed Scopus Google Scholar). some or of these mechanisms is presently It is however, that Bcl-2 family members and and that this the to apoptosis upon to stress Bcl-2 family members may by than For activation of BAD, at two serine and Bcl-2 or at the (16Wang H.-G. Rapp U.R. Reed J.C. Cell. 1996; 87: 629-638Abstract Full Text Full Text PDF PubMed Scopus (708) Google Scholar, 17Zha J. Harada H. Yang E. Jockel J. Korsmeyer S.J. Cell. 1996; 87: 619-628Abstract Full Text Full Text PDF PubMed Scopus (2253) Google Scholar). BAD to and and to a in a pro-apoptotic BAD Bcl-2 and is and by in the the anti-apoptotic of Recent studies that the kinase serine 136 of BAD in is S.R. H. X. S. H. Y. Cell. 1997; 91: Full Text Full Text PDF PubMed Scopus Google Scholar). The kinase for serine has not been The of appears to the mechanism by signals, and induction of apoptosis in cells J. Harada H. Yang E. Jockel J. Korsmeyer S.J. Cell. 1996; 87: 619-628Abstract Full Text Full Text PDF PubMed Scopus (2253) Google Scholar, L. M. C. R. G. 1997; PubMed Scopus Google Scholar). Further, the mechanism by pro-apoptotic Bcl-2 family members such as or signal apoptosis is BAD to serine and serine 136 are not present in these The present studies of BAD to ceramide signaling of apoptosis. that the pro-apoptotic Bcl-2 family member BAD ceramide-induced apoptosis onto COS-7 cells, are Ceramide signals apoptosis in BAD-expressing cells by a pathway involving sequentially of Ras (KSR)/ceramide-activated protein kinase Ras, and to BAD through Akt/PKB. These investigations that the availability of a single target, in this BAD, the mitogen-activated protein kinase cascade, is proliferative into a pro-apoptotic signaling COS-7 cells in and at in a as Y. B. S. S. S. M. H. Kolesnick R. Cell. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar). The and generated as Y. B. S. S. S. M. H. Kolesnick R. Cell. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar). The and by The and by The Ras and by A. The BAD and by The phosphatidylinositol and by COS-7 cells in and the of to the in for and to growth or amounts of BAD expression not by Ras or kinase For the cells or for to in growth or and cells by and cells. cells and in cells and cells for apoptosis by as F. M. S. R. R. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). Ras of Raf-1 activation by COS-7 cells in and as of KSR Raf-1 or as in The at for and Raf-1 the at Raf-1 by and to an kinase kinase-inactive as as Y. B. S. S. S. M. H. Kolesnick R. Cell. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar). by to the of ceramide on COS-7 cells in and as of or and in of and an kinase as as H. S.R. M.J. R. G.M. 1997; PubMed Scopus Google Scholar). COS-7 cells in and of or to or cells in of by either or by as Ceramide is a of apoptosis in a variety of cell systems (2Pena L.A. Fuks Z. Kolesnick R. Biochem. Pharmacol. 1997; 53: 615-621Crossref PubMed Scopus (220) Google Scholar, M.J. Obeid L.M. Hannun Y.A. Adv. Pharmacol. 1997; 41: 133-154Crossref PubMed Scopus (62) Google Scholar). In as as to apoptosis in COS-7 cells at B and data not Bcl-2 ceramide-induced apoptosis in systems (11Susin S.A. Zamzami N. Castedo M. Daugas E. Wang H.G. Geley S. Fassy F. Reed J.C. Kroemer G. J. Exp. Med. 1997; 186: 25-37Crossref PubMed Scopus (589) Google Scholar, 12Smyth M.J. Perry D.K. Zhang J. Poirier G.G. Hannun Y.A. Obeid L.M. Biochem. J. 1996; 316: 25-28Crossref PubMed Scopus (198) Google Scholar, 13Farschon D.M. Couture C. Mustelin T. Newmeyer D.D. J. Cell Biol. 1997; 137: 1117-1125Crossref PubMed Scopus (36) Google the of Bcl-2 family members in COS-7 cells. revealed of and in COS-7 cells, the pro-apoptotic Bcl-2 family member BAD not of BAD and apoptosis and For these COS-7 cells and to growth as as of the BAD expression apoptosis a of the and of the apoptosis to of the and the of of elevated apoptosis as as the and by ceramide BAD ceramide to apoptosis at to at of BAD than The for this of ceramide of the studies revealed that apoptosis to cells for BAD not BAD conferred ceramide-induced caspase activation onto COS-7 cells as a caspase not A. M. Y. M. Y.A. S.M. D.K. 1995; PubMed Scopus Google Scholar). in BAD-expressing cells not by to ceramide not expression of amounts of BAD ceramide to signal apoptosis in cells not a cell in ceramide induces a proliferative response S. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). For of as generated a of apoptosis and ceramide In to the of ceramide, the of not apoptosis These studies indicate an for the of the for the induction of apoptosis by The of ceramide to signal apoptosis in BAD-expressing cells as as apoptosis BAD and a The of the of a target for ceramide, on apoptosis. KSR in genetic as of Ras and either upstream or to Raf-1 K. Cell. 1995; Full Text PDF PubMed Scopus Google Scholar, M. M. Cell. 1995; Full Text PDF PubMed Scopus Google Scholar, M. G.M. Cell. 1995; Full Text PDF PubMed Scopus Google Scholar). Recent studies evidence that B. Zhang Y. S. S. S. Kolesnick R. 1995; PubMed Scopus Google Scholar) is the of KSR Y. B. S. S. S. M. H. Kolesnick R. Cell. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar). that on to signaling through the but not second in and in For the present the KSR is when in COS-7 cells Y. B. S. S. S. M. H. Kolesnick R. Cell. 1997; Full Text Full Text PDF PubMed Scopus Google or of to cells, ceramide, to initiate apoptosis However, BAD KSR to signal apoptosis in a at of KSR than of the kinase-inactive KSR generated by of for conserved to apoptosis In ceramide-induced apoptosis in BAD-expressing cells KSR BAD expression not ceramide signaling of apoptosis mediated by These studies of ceramide in a pathway to apoptosis. G. Rapp U.R. PubMed Scopus Google Scholar) to initiate apoptosis in COS-7 cells, but signaled apoptosis when BAD BAD Further, expression of G. Rapp U.R. PubMed Scopus Google Scholar) to signal apoptosis in BAD-expressing cells, the of BAD expression not studies to KSR and Raf-1 in signaling of the apoptotic response. that KSR to apoptosis, apoptosis the target for ceramide, as is upstream of Raf-1 in the signaling of apoptosis in BAD-expressing COS-7 cells. the specific of L. T. S.J. J. Biol. Chem. 1995; Full Text Full Text PDF PubMed Scopus Google ceramide-induced apoptosis data not In to ceramide-induced apoptosis not of KSR and in KSR KSR cells as in of BAD or and of KSR or or the and to a of of in cells as in These data independent in a COS-7 cells as in of BAD or and of KSR or or the and to a of of in cells as in These data independent a for Ras in apoptosis through BAD, studies Ras for For these Raf-1 the Ras for and Raf-1 and an kinase kinase-inactive as Y. B. S. S. S. M. H. Kolesnick R. Cell. 1997; Full Text Full Text PDF PubMed Scopus Google of KSR Raf-1 in Raf-1 This by L.A. G.M. Cell. Biol. PubMed Scopus Google but not by L.A. M.M. S. A. J. D. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar) or L.A. K. K.M. M.M. S. Cell. Biol. PubMed Scopus Google mutants the of and serve as when Raf-1 by the not These studies indicate that Ras is for activation of but not or ceramide-induced apoptosis in BAD-expressing COS-7 cells These indicate that Ras ceramide signaling of apoptosis through the of BAD on serine or in of BAD inactivation by as a and an anti-apoptotic (16Wang H.-G. Rapp U.R. Reed J.C. Cell. 1996; 87: 629-638Abstract Full Text Full Text PDF PubMed Scopus (708) Google Scholar, 17Zha J. Harada H. Yang E. Jockel J. Korsmeyer S.J. Cell. 1996; 87: 619-628Abstract Full Text Full Text PDF PubMed Scopus (2253) Google Scholar). of signals apoptosis. pro-apoptotic Bcl-2 family such as I. T. J.C. R. 1995; PubMed Scopus Google Scholar, T. R. C. 1995; PubMed Scopus Google Scholar) and Korsmeyer S.J. Cell. Full Text PDF PubMed Scopus Google not and are by signaling mechanisms than the to BAD, COS-7 cells and that and BAD, apoptosis when in COS-7 cells. In to BAD, however, and to ceramide signaling of apoptosis. These indicate that are in the ceramide-induced signaling to apoptosis in COS-7 cells. to that serine or 136 of BAD be to regulate this of apoptosis. elucidate the of these the serine mutants and J. Harada H. Yang E. Jockel J. Korsmeyer S.J. Cell. 1996; 87: 619-628Abstract Full Text Full Text PDF PubMed Scopus (2253) Google Scholar) in COS-7 cells and in apoptosis than of COS-7 cells of apoptosis, of apoptosis and of generated apoptosis not of serine 136 apoptosis in COS-7 cells of serine a It be that of of the BAD of For the of and to a of apoptosis of the of BAD, ceramide signaling of apoptosis, apoptosis but to ceramide not amounts of and at the of COS-7 cells, the of apoptosis in cells to that observed in BAD-expressing cells ceramide not These studies the serine 136 of BAD as for ceramide signaling of apoptosis through Further, that ceramide dephosphorylation of BAD to for signaling of apoptosis. this the of BAD and ceramide to has been in cells J. Harada H. Yang E. Jockel J. Korsmeyer S.J. Cell. 1996; 87: 619-628Abstract Full Text Full Text PDF PubMed Scopus (2253) Google BAD as a in COS-7 cells, as by an The is on serine 136 a specific for S.R. H. X. S. H. Y. Cell. 1997; 91: Full Text Full Text PDF PubMed Scopus Google this not Further, of cells ceramide in of the of BAD by the of the in and of the by and S.R. H. X. S. H. Y. Cell. 1997; 91: Full Text Full Text PDF PubMed Scopus Google the be on a gel as two by of by The in of ceramide not of COS-7 cells apoptosis, dephosphorylation of BAD in response to ceramide, that ceramide signals this via the this and not Recent investigations that apoptosis by in cell Cell. 1997; Full Text Full Text PDF PubMed Scopus Google BAD at serine apoptosis S.R. H. X. S. H. Y. Cell. 1997; 91: Full Text Full Text PDF PubMed Scopus Google Scholar, L. M. C. R. G. 1997; PubMed Scopus Google Scholar). the upstream signaling via ceramide into the COS-7 cells and an kinase as a that ceramide to a in This as as ceramide not the upstream pathway by ceramide at the of the or at a of the F. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google or a generated by the of to the A. 1995; PubMed Scopus Google BAD in COS-7 cells. ceramide-induced apoptosis, to expression of by of in a in apoptosis, not the in apoptosis in response to ceramide not These studies support the that ceramide signals apoptosis by of The present studies signaling through Ras and Raf-1 to apoptosis via dephosphorylation of the pro-apoptotic Bcl-2 family member COS-7 cells, BAD, and cells, BAD, to to ceramide apoptosis, expression of small amounts of BAD this BAD apoptosis onto a ceramide target, and KSR or Raf-1 ceramide signaling of apoptosis through of these and revealed that KSR is upstream of Raf-1 in the to apoptosis. these studies the genetic studies K. Cell. 1995; Full Text PDF PubMed Scopus Google Scholar, M. M. Cell. 1995; Full Text PDF PubMed Scopus Google Scholar, M. G.M. Cell. 1995; Full Text PDF PubMed Scopus Google Scholar, J. Cell. 1995; Full Text PDF PubMed Scopus Google not to KSR upstream or to Ras is for signaling of apoptosis through that the of these upstream is proliferation or apoptosis is the final outcome. role for BAD in this by mutation at serine abrogated the pro-apoptotic of ceramide in COS-7 cells. These studies support the that the availability of a single target, in this BAD, a proliferative signaling to an apoptotic response. These are the regulation of ceramide signaling of apoptosis through Bcl-2 family Kroemer and D. Geley S. Hirsch T. Castedo M. Marchetti P. Macho A. R. Kroemer G. Res. 1997; Google Scholar) have that ceramide signals of and generation of in these are transcriptionally support for this is studies in a cell-free system, that cells ceramide analogs the to in to apoptosis of (15Susin S. Zamzami N. Larochette N. Dallaporta B. Marzo I. Brenner C. Hirsch T. Petit P. Geuskens M. Kroemer G. Exp. Cell Res. 1997; 236: 397-403Crossref PubMed Scopus (75) Google Scholar). Ceramide-induced in cells, and the cell-free system by Bcl-2 (14Castedo M. Hirsch T. Susin S.A. Zamzami N. Marchetti P. Macho A. Kroemer G. J. Immunol. 1996; 157: 512-521PubMed Google Scholar, S. Zamzami N. Larochette N. Dallaporta B. Marzo I. Brenner C. Hirsch T. Petit P. Geuskens M. Kroemer G. Exp. Cell Res. 1997; 236: 397-403Crossref PubMed Scopus (75) Google Scholar, D. Geley S. Hirsch T. Castedo M. Marchetti P. Macho A. R. Kroemer G. Res. 1997; Google Scholar). ceramide signaling of apoptosis in BAD-expressing COS-7 cells not by the the and BAD not protein Ceramide signaling of BAD dephosphorylation appears to through inactivation of Akt/PKB. Ceramide in BAD-expressing cells, and but not ceramide-induced apoptosis. This latter that is that activation of a this have observed that ceramide in by a mechanism independent of that this pathway may in cells than COS-7 cells. J. It be that BAD has a S. M. Zhang X. D. Wang H.G. A. G. S. Reed J.C. J. 1998; Google Scholar) ceramide-induced apoptosis is (2Pena L.A. Fuks Z. Kolesnick R. Biochem. Pharmacol. 1997; 53: 615-621Crossref PubMed Scopus (220) Google Scholar, M.J. Obeid L.M. Hannun Y.A. Adv. Pharmacol. 1997; 41: 133-154Crossref PubMed Scopus (62) Google Scholar). this mechanism is not to mediate of ceramide signaling of apoptosis, Bcl-2 is that the This that ceramide, Cell. 1997; Full Text Full Text PDF PubMed Scopus Google must to effectors of the apoptotic and are not as ceramide-induced apoptosis in COS-7 cells. In activation of the Ras, Raf-1 and pathway is and anti-apoptotic Z. M. J. R. M. 1995; PubMed Scopus Google Scholar, O. G. B. Spiegel S. 1996; PubMed Scopus Google Scholar). However, investigations have begun to in Ras signals apoptosis. Gulbins and E. R. A. Martin S. T. G. G. C. Lang F. 1995; Full Text PDF PubMed Scopus Google Scholar, E. K.M. Brenner B. K. Linderkamp O. Lang F. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar) that activation of in cells, and that mutants of Ras E. J. A. E. I. Proc. Natl. Acad. Sci. U. S. A. 1997; PubMed Scopus Google Scholar) this to a for in death of a cell J.C. S.M. A. EMBO J. 1996; PubMed Scopus Google Scholar) in cells the present studies not or Ras is for ceramide-induced ceramide is of Ras, by an in cells E. R. A. Martin S. T. G. G. C. Lang F. 1995; Full Text PDF PubMed Scopus Google Scholar, E. K.M. Brenner B. K. Linderkamp O. Lang F. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar). Ras signaling appears as the cell apoptosis or may depending on the genetic and For and of apoptosis of K. Cell. Biol. 1997; PubMed Scopus Google Scholar). of cells to apoptosis protein kinase or 1995; Google Scholar). Further, Ras apoptosis in cells activation of by of a of Wang Wang 1997; PubMed Scopus Google Scholar). In some the to may gene For expression of small amounts of Raf-1 of cells expression induction of apoptosis D. S. 1997; PubMed Scopus Google Scholar). study by and A. P. E. C. P. J. G. 1997; PubMed Scopus Google Scholar) the of of the Ras effector systems mediate apoptosis in a of In in is through an of apoptosis. of mutants of these that apoptosis mediated through on apoptosis, and signaled These studies that the outcome of signaling through Ras be at in by the of these effector In some of the apoptosis, apoptosis, inhibitable by Bcl-2 E. J. A. E. I. Proc. Natl. Acad. Sci. U. S. A. 1997; PubMed Scopus Google Scholar, 1995; Google Scholar, M. D. T. S. 1997; PubMed Scopus Google the that be The present studies investigations into ceramide signaling through B. Zhang Y. S. S. S. Kolesnick R. 1995; PubMed Scopus Google Scholar, S. Kolesnick Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, R. Kolesnick J. Biol. Chem. Full Text PDF PubMed Google Scholar, J. S. Yang Z. Kolesnick J. Biol. Chem. Full Text PDF PubMed Google Scholar, S. Kolesnick PubMed Scopus Google Scholar). on the of ceramide but not to KSR in and in and a for the of Raf-1 for KSR that KSR and are Y. B. S. S. S. M. H. Kolesnick R. Cell. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar). In the present KSR the of ceramide to signal apoptosis through Ras and and that the kinase of KSR Further, ceramide-induced apoptosis through BAD, the role of KSR as a target for have at as to the mechanism by KSR and M. G.M. D.K. 1996; PubMed Scopus Google and and H. K. A. Biol. 1997; Full Text Full Text PDF PubMed Scopus Google that KSR to and signaling through the This is as for and although is as to the kinase domain of KSR is M. A. Spiegel S. G.M. D.K. Proc. Natl. Acad. Sci. U. S. A. 1997; PubMed Scopus Google Scholar). In and G. L. Biol. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar) as as and A. C. D. G. J. A. Biol. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar) that KSR to and signaling through and and Perhaps the in these data induction of apoptosis. and G.M. 1998; Google Scholar) have recently that of at gene induces of cells in and of expression apoptosis. and as of mutants and In some of the systems apoptosis may for the occurs of KSR into COS-7 cells, for a in of KSR The present studies of transcriptionally independent apoptosis in response to ceramide signaling through and the investigations have of proliferative and apoptotic signaling E. J. A. E. I. Proc. Natl. Acad. Sci. U. S. A. 1997; PubMed Scopus Google K. Cell. Biol. 1997; PubMed Scopus Google Scholar, A. P. E. C. P. J. G. 1997; PubMed Scopus Google Scholar). The present study that the availability of a single target may dictate the final outcome.