Protein Kinase Cι Activity Is Necessary for Bcr-Abl-mediated Resistance to Drug-induced Apoptosis

Abstract

K562 chronic myelogenous leukemia cells are highly resistant to chemotherapeutic drugs, such as taxol, that induce cell death by apoptosis. This resistance is mediated by the chimeric tyrosine kinase oncogene Bcr-Abl. However, little is known about the mechanism by which Bcr-Abl protects K562 cells from apoptosis. We recently demonstrated that expression of PKCι is necessary for the resistance of K562 cells to taxol-induced apoptosis (Murray, N. R., and Fields, A. P. (1997) J. Biol. Chem. 272, 27521–27524). We now demonstrate that treatment of K562 cells with taxol leads to sustained activation of PKCι. In contrast, Bcr-Abl-negative HL60 myeloid leukemia cells, which are sensitive to taxol-induced apoptosis, do not exhibit sustained PKCι activation in response to taxol. Treatment of K562 cells with tyrphostin AG957, a selective Bcr-Abl inhibitor, blocks taxol-induced PKCι activation and sensitizes these cells to taxol-induced apoptosis, indicating that PKCι is a relevant downstream target of Bcr-Abl-mediated resistance. Furthermore, expression of constitutively active PKCι by adenovirus-mediated gene transfer rescues AG957-treated K562 cells from taxol-induced apoptosis. Taken together, these results demonstrate that both Bcr-Abl and PKCι activity are necessary for apoptotic resistance in K562 cells. Furthermore, they identify PKCι as a critical downstream target of Bcr-Abl that is sufficient to mediate the anti-apoptotic effects of Bcr-Abl. K562 chronic myelogenous leukemia cells are highly resistant to chemotherapeutic drugs, such as taxol, that induce cell death by apoptosis. This resistance is mediated by the chimeric tyrosine kinase oncogene Bcr-Abl. However, little is known about the mechanism by which Bcr-Abl protects K562 cells from apoptosis. We recently demonstrated that expression of PKCι is necessary for the resistance of K562 cells to taxol-induced apoptosis (Murray, N. R., and Fields, A. P. (1997) J. Biol. Chem. 272, 27521–27524). We now demonstrate that treatment of K562 cells with taxol leads to sustained activation of PKCι. In contrast, Bcr-Abl-negative HL60 myeloid leukemia cells, which are sensitive to taxol-induced apoptosis, do not exhibit sustained PKCι activation in response to taxol. Treatment of K562 cells with tyrphostin AG957, a selective Bcr-Abl inhibitor, blocks taxol-induced PKCι activation and sensitizes these cells to taxol-induced apoptosis, indicating that PKCι is a relevant downstream target of Bcr-Abl-mediated resistance. Furthermore, expression of constitutively active PKCι by adenovirus-mediated gene transfer rescues AG957-treated K562 cells from taxol-induced apoptosis. Taken together, these results demonstrate that both Bcr-Abl and PKCι activity are necessary for apoptotic resistance in K562 cells. Furthermore, they identify PKCι as a critical downstream target of Bcr-Abl that is sufficient to mediate the anti-apoptotic effects of Bcr-Abl. Leukemia is a malignant disease of the bone marrow that is the leading cause of cancer death in children and adults under the age of 35 (1Landis S.H. Murray T. Parker S.L. Tong T. Bolden S. Wingo P.A. Cancer J. Clin. 1998; 48: 6-30Crossref PubMed Scopus (2456) Google Scholar). Leukemias arise in hematopoietic progenitor cells and are characterized by impaired or blocked differentiation, uncontrolled proliferation, and resistance to apoptosis (2Hiddemann W. Griesinger F. Curr. Opin. Oncol. 1993; 5: 3-25PubMed Google Scholar). Many leukemias can be effectively treated using one or combinations of chemotherapeutic agents, such as taxol, that induce apoptosis (reviewed in Ref. 3Kerr J.F. Winterford C.M. Harmon B.V. Cancer. 1994; 73: 2013-2026Crossref PubMed Scopus (2167) Google Scholar). A notable exception is chronic myelogenous leukemia (CML), 1The abbreviations used are: CML, chronic myelogenous leukemia; PKC, protein kinase C; PI, phosphatidylinositol; GFP, green fluorescent protein. which is highly resistant to most commonly utilized chemotherapeutic drugs, including taxol (4Green D.R. Bissonnette R.P. Cotter T.G. Import. Adv. Oncol. 1994; 1994: 37-52Google Scholar, 5Bedi A. Barber J.P. Bedi G.C. el-Deiry W.S. Sidransky D. Vala M.S. Akhtar A.J. Hilton J. Jones R.J. Blood. 1995; 86: 1148-1158Crossref PubMed Google Scholar). CML cells, such as K562 cells, are characterized by the presence of the Philadelphia chromosome, which results from a reciprocal translocation involving chromosome 9 and chromosome 22 (6Nowell P.C. Hungerford D.A. J. Natl. Cancer Inst. 1960; 25: 85-89PubMed Google Scholar,7Rowley J.D. Nature. 1973; 243: 290-293Crossref PubMed Scopus (3403) Google Scholar). In K562 cells, the Abl tyrosine kinase gene on chromosome 9 is translocated into the breakpoint cluster region (bcr) region on chromosome 22 producing a chimeric gene whose product, Bcr-Abl, expresses disregulated Abl tyrosine kinase activity (8Kurzrock R. Gutterman J.U. Talpaz M. New Engl. J. Med. 1988; 319: 990-998Crossref PubMed Scopus (735) Google Scholar). The Bcr-Abl gene is the transforming activity responsible for CML (9Daley G.Q. Van Etten R.A. Baltimore D. Science. 1990; 247: 824-830Crossref PubMed Scopus (1929) Google Scholar), and the resistance of K562 cells to drug-induced apoptosis is conferred by Bcr-Abl (5Bedi A. Barber J.P. Bedi G.C. el-Deiry W.S. Sidransky D. Vala M.S. Akhtar A.J. Hilton J. Jones R.J. Blood. 1995; 86: 1148-1158Crossref PubMed Google Scholar, 10Cortez D. Kadlec L. Pendergast A.M. Mol. Cell. Biol. 1995; 15: 5531-5541Crossref PubMed Scopus (268) Google Scholar, 11McGahon A. Bissonnette R. Schmitt M. Cotter K.M. Green D.R. Cotter T.G. Blood. 1994; 83: 1179-1187Crossref PubMed Google Scholar). Thus, inhibition of Bcr-Abl tyrosine kinase activity using selective inhibitors (12Anafi M. Gazit A. Zehavi A. Ben-Neriah Y. Levitzki A. Blood. 1993; 82: 3524-3529Crossref PubMed Google Scholar, 13Riordan F.A. Bravery C.A. Mengubas K. Ray N. Borthwick N.J. Akbar A.N. Hart S.M. Hoffbrand A.V. Mehta A.B. Wickremasinghe R.G. Oncogene. 1998; 16: 1533-1542Crossref PubMed Scopus (28) Google Scholar, 14Deora A.B. Miranda M.B. Rao S.G. Tumorigenesis. 1997; 83: 756-761PubMed Google Scholar, 15Beran M. Cao X. Estrov Z. Jeha S. Jin G. O'Brien S. Talpaz M. Arlinghaus R.B. Lydon N.B. Kantarjian H. Clin. Cancer Res. 1998; 4: 1661-1672PubMed Google Scholar) or of Bcr-Abl expression using antisense oligonucleotides (16McGahon A.J. Brown D.B. Martin S.J. Amarante-Mendes G.P. Cotter T.G. Cohen G.M. Green D.R. Cell Growth Differ. 1997; 4: 95-103Crossref Scopus (43) Google Scholar, 17Rowley P.T. Keng P.C. Kosciolek B.A. Leuk. Res. 1996; 20: 473-480Crossref PubMed Scopus (36) Google Scholar) leads to increased sensitivity of K562 cells to apoptosis induced by many drugs including taxol. Despite the importance of Bcr-Abl in resistance to drug-induced apoptosis, relatively little is known about the molecular mechanisms by which Bcr-Abl exerts its anti-apoptotic effects (18Amarante-Mendes G.P. McGahon A.J. Nishioka W.K. Afar D.E.H. Witte O.N. Green D.R. Oncogene. 1998; 16: 1383-1390Crossref PubMed Scopus (200) Google Scholar). Recently, we demonstrated that expression of the atypical PKC isozyme, PKCι, is important for resistance to taxol-induced apoptosis (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar). We have now further defined the role of PKCι in this process. We find that treatment of K562 cells with taxol leads to potent and sustained activation of PKCι. In contrast, treatment of HL60 cells, which are sensitive to taxol-induced apoptosis, does not lead to appreciable activation of PKCι. Furthermore, inhibition of Bcr-Abl using the highly selective inhibitor tyrphostin AG957 blocks PKCι activation and sensitizes K562 cells to taxol-induced apoptosis. Tyrphostin AG957-treated cells can be rescued from taxol-induced apoptosis by expression of constitutively active PKCι. These data, along with our previous results (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar), demonstrate that PKCι activation is both necessary and sufficient to mediated the anti-apoptotic effects of Bcr-Abl. Human K562 chronic myelogenous leukemia cells and HL60 promyelocytic leukemia cells were maintained in suspension culture as described previously (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar). Cells at a density of 2 × 105 cells/ml were treated with paclitaxel (taxol) at the concentrations and for the time periods indicated in the figure legends. In some cases, the Bcr-Abl-selective inhibitor tyrphostin AG957 or the PI 3-kinase inhibitors wortmannin or LY 294002 were added to K562 cell cultures at the concentrations indicated in the figure legends prior to treatment with taxol. Tyrphostin AG957 is a highly selective Bcr-Abl inhibitor that blocks Bcr-Abl-mediated cell growth and resistance to apoptosis in K562 cells (12Anafi M. Gazit A. Zehavi A. Ben-Neriah Y. Levitzki A. Blood. 1993; 82: 3524-3529Crossref PubMed Google Scholar, 20Kaur G. Sausville E.A. Anticancer Drugs. 1996; 7: 815-824Crossref PubMed Scopus (17) Google Scholar). Apoptosis was assessed by the presence of apoptotic bodies or chromosomal condensation and by trypan blue exclusion as described previously (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar). These morphological manifestations of apoptosis correlate directly with the apoptotic DNA fragmentation induced by taxol and provide a convenient method to quantitate the apoptotic response (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar). For assessment of nuclear DNA morphology, 1 × 105 cells were fixed and permeablized with 1% paraformaldehyde phosphate-buffered saline for 15 min at 37 °C, stained with 4′,6-diamidino-2-phenylindole for 5 min, and observed under phase-fluorescence optics. A minimum of 500 cells for each treatment were scored for the presence of apoptotic bodies. K562 and HL60 cell extracts were lysed directly into SDS sample buffer and subjected to SDS-polyacrylamide gel electrophoresis and immunoblot analysis for PKCι as described previously (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar). We have previously demonstrated that K562 cells express PKCι but no detectable PKCζ (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar). Protein quantitation was carried out using BCA protein assay reagent (Pierce). Antigen/antibody complexes were detected using ECL (Amersham Pharmacia Biotech). For immunoprecipitation kinase assay of PKCι, K562 and HL60 cells were lysed in RIPA lysis buffer (150 mm NaCl, 1% Nonidet P-40, 0.5% deoxycholate, 0.1% SDS, 50 mm Tris, pH 8.0) at 4 °C in the presence of 1 mm phenylmethylsulfonyl fluoride and 20 μg/ml leupeptin. Lysates were precleared by incubation with 50 μl of protein A-Sepharose beads (Sigma) for 1 h at 4 °C. PKCι antibody was first bound to 50 μl of protein A-Sepharose at 4 °C for 1 h prior to addition to clarified lysates and further incubation at 4 °C for 1 h. Immunoprecipitates were washed once with RIPA buffer and once with kinase assay buffer (50 mm Tris, pH 7.5, 10 mmMgCl2, 0.5 mm EGTA, 0.1 mmCaCl2) to remove unbound proteins. Washed beads were resuspended in kinase buffer containing 40 μg/ml phosphatidylserine, 10 μg of histone substrate, 10 μm ATP, and 10 μCi of [γ-32P]ATP. The reactions were incubated at room temperature for 30 min and terminated by addition of SDS buffer and boiling prior to SDS-polyacrylamide gel electrophoresis and autoradiography. Constitutively active PKCι was generated by PCR amplification of a fragment containing an A120E substitution within the pseudosubstrate domain of human PKCι. Such a mutation has previously been shown to induce constitutive activity in PKCι (21Diaz-Meco M.T. Municio M.M. Frutos S. Sanchez P. Lozano J. Sanz L. Moscat J. Cell. 1996; 86: 777-786Abstract Full Text Full Text PDF PubMed Scopus (328) Google Scholar). TheEcoRV/NdeI-digested fragment was subcloned into similarly digested full-length PKCι cDNA and sequenced to confirm the presence of this single mutation. PKCιA120E (designated caPKCι) was then subcloned into pXCX.CMV, a transfer vector based on pXCX2 (22Spessot R. Inchley K. Hupel T.M. Bacchetti S. Virology. 1989; 168: 378-387Crossref PubMed Scopus (41) Google Scholar) containing the expression cassette from pRcCMV (Stratagene). Recombinant adenoviruses containing either the green fluorescent protein (GFP) (a kind gift from Phil Poronik, University of Sydney, NSW, Australia) or constitutively active PKCι were produced by homologous recombination with pJM17 (a circular form of the adenovirus genome) in HEK 293 cells as described (23Graham F.L. Prevec L. Murray E.J. Methods in Molecular Biology: Manipulation of Adenovirus Vectors Gene Transfer and Expression Protocols. 7. Humana Press Scholar). were and characterized by lysates were generated by of HEK 293 cells as described (23Graham F.L. Prevec L. Murray E.J. Methods in Molecular Biology: Manipulation of Adenovirus Vectors Gene Transfer and Expression Protocols. 7. Humana Press Scholar). lysates of were used to K562 cells. was into K562 cells using the reagent cells were treated with taxol AG957 for at which time cells were for apoptosis as described was as by for expression was by immunoprecipitation kinase assay of cell lysates from cells h as described Recently, we demonstrated that PKCι expression is important for resistance of K562 cells to apoptotic including taxol (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar). of PKCι leads to resistance to taxol-induced apoptosis, antisense inhibition of PKCι expression leads to to taxol-induced apoptosis (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar). These results that in PKCι expression activity a critical role in the of K562 cells to apoptotic the of PKCι expression is a of resistance to drug-induced apoptosis, we human cell that in sensitivity to taxol-induced apoptosis. HL60 cells are sensitive to taxol, apoptosis such that in the presence of taxol, of HL60 cells are apoptotic h In contrast, K562 cells are highly resistant to taxol-induced apoptosis little or no apoptosis h in to taxol this in resistance to taxol-induced apoptosis with PKCι both cell were treated with taxol for to 50 and cell lysates were subjected to immunoblot analysis for PKCι as described previously (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar) these cell express of PKCι A and Furthermore, PKCι do not in response to taxol-induced apoptotic that in resistance to apoptosis are not to in PKCι we PKCι activity in K562 and HL60 cells taxol-induced apoptotic 2 Cells were treated with taxol and PKCι from cell lysates at the indicated and for PKCι activity as described under analysis demonstrated that of PKCι is by this antibody not PKCι activity is in K562 and HL60 cells prior to to taxol, with the of PKCι expression in these cells. HL60 cells are treated with taxol, little in PKCι activity is observed the first by which time of HL60 cells are PKCι activity to by h. This of PKCι activity is not to the of PKCι do not this time 2 In contrast, PKCι activity in K562 cells is induced in response to taxol 2 PKCι activity is at at and is sustained the h. of the a in PKCι activity by h. These demonstrate that PKCι is in response to apoptotic in K562 cells but not HL60 cells, that sustained PKCι activation be important in the resistance of K562 cells to taxol-induced apoptosis. The Bcr-Abl tyrosine kinase is the oncogene responsible for CML and is highly in K562 cells but is from HL60 cells. Bcr-Abl is for both and resistance to drug-induced apoptosis of K562 cells (5Bedi A. Barber J.P. Bedi G.C. el-Deiry W.S. Sidransky D. Vala M.S. Akhtar A.J. Hilton J. Jones R.J. Blood. 1995; 86: 1148-1158Crossref PubMed Google Scholar, 10Cortez D. Kadlec L. Pendergast A.M. Mol. Cell. Biol. 1995; 15: 5531-5541Crossref PubMed Scopus (268) Google Scholar, 11McGahon A. Bissonnette R. Schmitt M. Cotter K.M. Green D.R. Cotter T.G. Blood. 1994; 83: 1179-1187Crossref PubMed Google Scholar). However, the downstream of Bcr-Abl critical for the resistance are not the of PKCι activation in taxol we taxol-induced PKCι activation in K562 cells was Bcr-Abl kinase Furthermore, Bcr-Abl can directly and PI 3-kinase A. K. K. K. 1994; Google Scholar, M. R. K. N. G. J.D. Oncogene. 1996; Google Scholar) to PI which can in PKCι K. R. S. Nishioka N. J. K. Y. S. K. S. A. S. J. 1996; 15: PubMed Scopus Google Scholar), we PI 3-kinase activity was for taxol-induced PKCι K562 cells were treated with in the presence or of the Bcr-Abl inhibitor tyrphostin AG957 (12Anafi M. Gazit A. Zehavi A. Ben-Neriah Y. Levitzki A. Blood. 1993; 82: 3524-3529Crossref PubMed Google Scholar, 20Kaur G. Sausville E.A. Anticancer Drugs. 1996; 7: 815-824Crossref PubMed Scopus (17) Google Scholar), or the PI 3-kinase inhibitors wortmannin T. L. Y. M. J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar) or Brown J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar) for h. PKCι activity was by immunoprecipitation kinase treatment leads to potent activation of PKCι the activity in the of taxol However, tyrphostin AG957 taxol-induced PKCι wortmannin PKCι activation the that they were used at concentrations for wortmannin T. L. Y. M. J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar) and μm for Brown J. Biol. Chem. 1994; Full Text PDF PubMed Google Tyrphostin inhibition of taxol-induced PKCι activation is and is at 30 μm with the of the that Bcr-Abl activity (12Anafi M. Gazit A. Zehavi A. Ben-Neriah Y. Levitzki A. Blood. 1993; 82: 3524-3529Crossref PubMed Google Scholar, 20Kaur G. Sausville E.A. Anticancer Drugs. 1996; 7: 815-824Crossref PubMed Scopus (17) Google Scholar). Furthermore, tyrphostin AG957 leads to of K562 cells to taxol-induced apoptosis with its to PKCι activation wortmannin on taxol-induced apoptosis, with the of inhibition of PKCι observed with these PI 3-kinase Taken together, these that Bcr-Abl is an of taxol-induced PKCι that PKCι activation is important for resistance of K562 cells to taxol, and that Bcr-Abl-mediated PKCι activation does not PI 3-kinase PKCι activity is sufficient to K562 cells from taxol-induced apoptosis, we used adenovirus gene transfer to express constitutively active PKCι in K562 cells. Constitutively active PKCι adenovirus was produced by mutation of to in the pseudosubstrate region of PKCι as described under This mutation has been demonstrated to lead to constitutive activation of PKCι (21Diaz-Meco M.T. Municio M.M. Frutos S. Sanchez P. Lozano J. Sanz L. Moscat J. Cell. 1996; 86: 777-786Abstract Full Text Full Text PDF PubMed Scopus (328) Google Scholar). K562 cells were with either or adenovirus for h. demonstrated that of cells are under these kinase demonstrate that cells PKCι activity with cells In contrast, cells with of PKCι cells were treated with taxol and tyrphostin AG957 for and the of apoptosis was assessed 4 taxol tyrphostin AG957 induced apoptosis in or K562 cells. However, in the presence of tyrphostin AG957, taxol induced apoptosis in cells. In contrast, expression of blocked taxol-induced apoptosis in tyrphostin cells. These results demonstrate that PKCι is a downstream target of Bcr-Abl and that PKCι activity is sufficient to mediate the anti-apoptotic effects of Bcr-Abl. PKC is a of in the of proliferation, differentiation, and apoptosis (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar, N.R. Fields A.P. Parker L. Protein Molecular R. G. Scholar, J. G. Mol. 1997; PubMed Scopus Google Scholar, N.R. G.P. Fields A.P. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar, B.A. Fields A.P. J. Cell PubMed Google Scholar). We have characterized the of the PKC in K562 cells, and selective and inhibition of expression of each (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar, N.R. G.P. Fields A.P. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar, B.A. Fields A.P. J. Cell PubMed Google Scholar, B.A. C.A. Fields A.P. J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar). of leads to gene and increased sensitivity to differentiation, inhibition of expression blocks differentiation, that a role in this N.R. G.P. Fields A.P. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). In contrast, of leads to a in and resistance to N.R. G.P. Fields A.P. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). of expression blocks proliferation, that is for this N.R. G.P. Fields A.P. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). The effects of are at in the cell translocation and activation of the at the B.A. Fields A.P. J. Cell PubMed Google Scholar, B.A. C.A. Fields A.P. J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar). directly the nuclear protein at in nuclear and cell the B.A. C.A. Fields A.P. J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar, Fields A.P. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar, B.A. Fields A.P. J. Biol. Chem. 1993; Full Text PDF PubMed Google Scholar). PKCι no role in K562 cell or (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar). of PKCι leads to resistance to drug-induced apoptosis, and antisense inhibition of PKCι expression leads to increased sensitivity to apoptosis, indicating an important role in resistance to drug-induced apoptosis (19Murray N.R. Fields A.P. J. Biol. Chem. 1997; 272: 27521-27524Abstract Full Text Full Text PDF PubMed Scopus (158) Google Scholar). PKC can cells apoptosis (21Diaz-Meco M.T. Municio M.M. Frutos S. Sanchez P. Lozano J. Sanz L. Moscat J. Cell. 1996; 86: 777-786Abstract Full Text Full Text PDF PubMed Scopus (328) Google Scholar), indicating that atypical a role in cell and from apoptotic However, the mechanism by which PKCι protects cells from apoptosis is not In cells, atypical PKC activity is by prior to the of apoptosis, that atypical PKC activity is for cell (21Diaz-Meco M.T. Municio M.M. Frutos S. Sanchez P. Lozano J. Sanz L. Moscat J. Cell. 1996; 86: 777-786Abstract Full Text Full Text PDF PubMed Scopus (328) Google Scholar). In this and our previous we provide that PKCι activity is both necessary and sufficient for resistance of K562 cells to drug-induced apoptosis. results with PKCι are of described previously for Bcr-Abl, whose expression and tyrosine kinase activity is for resistance of K562 cells to drug-induced apoptosis (5Bedi A. Barber J.P. Bedi G.C. el-Deiry W.S. Sidransky D. Vala M.S. Akhtar A.J. Hilton J. Jones R.J. Blood. 1995; 86: 1148-1158Crossref PubMed Google Scholar, 10Cortez D. Kadlec L. Pendergast A.M. Mol. Cell. Biol. 1995; 15: 5531-5541Crossref PubMed Scopus (268) Google Scholar, 11McGahon A. Bissonnette R. Schmitt M. Cotter K.M. Green D.R. Cotter T.G. Blood. 1994; 83: 1179-1187Crossref PubMed Google Scholar, M. Gazit A. Zehavi A. Ben-Neriah Y. Levitzki A. Blood. 1993; 82: 3524-3529Crossref PubMed Google Scholar, 13Riordan F.A. Bravery C.A. Mengubas K. Ray N. Borthwick N.J. Akbar A.N. Hart S.M. Hoffbrand A.V. Mehta A.B. Wickremasinghe R.G. Oncogene. 1998; 16: 1533-1542Crossref PubMed Scopus (28) Google Scholar, 14Deora A.B. Miranda M.B. Rao S.G. Tumorigenesis. 1997; 83: 756-761PubMed Google Scholar, 15Beran M. Cao X. Estrov Z. Jeha S. Jin G. O'Brien S. Talpaz M. Arlinghaus R.B. Lydon N.B. Kantarjian H. Clin. Cancer Res. 1998; 4: 1661-1672PubMed Google Scholar). both HL60 and K562 cells express of PKCι, the of PKCι activity in these cells apoptotic is apoptotic sustained activation of PKCι is observed in K562 cells but not HL60 cells, that K562 cells an of PKCι activity that is either not or is in HL60 cells. Furthermore, we find that inhibition of Bcr-Abl blocks taxol-induced PKCι activation and apoptosis in K562 cells, that Bcr-Abl is for taxol-induced PKCι of our results is the that PI 3-kinase activity is not for Bcr-Abl-mediated PKCι activation or from taxol-induced apoptosis. PI can atypical PKC A. K. K. K. 1994; Google Scholar). results do not of the PI 3-kinase in the activation of PKCι in response to Bcr-Abl is a characterized whose transforming is to its anti-apoptotic of Bcr-Abl activity with the tyrosine kinase inhibitors or M. Cao X. Estrov Z. Jeha S. Jin G. O'Brien S. Talpaz M. Arlinghaus R.B. Lydon N.B. Kantarjian H. Clin. Cancer Res. 1998; 4: 1661-1672PubMed Google Scholar) or Bcr-Abl expression with antisense oligonucleotides Bcr-Abl (16McGahon A.J. Brown D.B. Martin S.J. Amarante-Mendes G.P. Cotter T.G. Cohen G.M. Green D.R. Cell Growth Differ. 1997; 4: 95-103Crossref Scopus (43) Google Scholar, 17Rowley P.T. Keng P.C. Kosciolek B.A. Leuk. Res. 1996; 20: 473-480Crossref PubMed Scopus (36) Google Scholar) sensitizes K562 cells to apoptosis. results using the highly selective Bcr-Abl inhibitor tyrphostin AG957 are with these and demonstrate that PKCι activation is a downstream target of Bcr-Abl necessary for its anti-apoptotic on the molecular mechanism by which Bcr-Abl PKCι and on of the downstream of PKCι that mediate its anti-apoptotic