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  • Original Paper
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Apoptosis of hematopoietic cells induced by growth factor withdrawal is associated with caspase-9 mediated cleavage of Raf-1

Abstract

The Raf-1 serine/threonine kinase is a key protein that is implicated in the transmission of many growth and cell survival signals. In the present study we demonstrate that apoptosis of hematopoietic cells induced by IL-3-deprivation is associated with the cleavage of Raf-1, resulting in the separation of the N-terminal regulatory domain and the C-terminal kinase domain. Raf-1 cleavage specifically occurs upon triggering of the mitochondrial death pathway, and coincides with the activation of specific caspases. Moreover, Bcl-2 overexpression or treatment with the caspase inhibitor z-VAD.fmk completely prevented Raf-1 cleavage, whereas caspase inhibition by treatment of cells with Ac-DEVD.fmk or z-IETD.fmk, or CrmA overexpression had no effect. Furthermore, in vitro cleavage studies indicate that caspase-9, which is the apical protease in the mitochondrial death pathway, is able to cleave Raf-1 at position D279. Cell fractionation studies showed that the Raf-1 C-terminal fragment that is generated upon IL-3 withdrawal is localized predominantly to the mitochondria. In addition, constitutive expression of this C-terminal Raf-1 fragment fused to a mitochondrial targeting sequence in Ba/F3 pre-B cells significantly delays apoptosis induced by IL-3 withdrawal. These results suggest an important role for caspase-9 mediated cleavage of Raf-1 in the negative feedback regulation of hematopoietic cell apoptosis induced by growth factor withdrawal.

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References

  • Alavi A, Hood JD, Frausto R, Stupack DG and Cheresh DA . (2003). Science, 301, 94–96.

  • Boatright KM, Renatus M, Scott FL, Sperandio S, Shin H, Pedersen IM, Ricci JE, Edris WA, Sutherlin DP, Green DR and Salvesen GS . (2003). Mol. Cell, 11, 529–541.

  • Boatright KM and Salvesen GS . (2003). Curr. Opin. Cell Biol., 15, 725–731.

  • Bojes HK, Feng X, Kehrer JP and Cohen GM . (1999). Cell Death Differ., 6, 61–70.

  • Bokoch GM . (1998). Cell Death Differ., 5, 637–645.

  • Boldin MP, Goncharov TM, Goltsev YV and Wallach D . (1996). Cell, 85, 803–815.

  • Chen J, Fujii K, Zhang L, Roberts T and Fu H . (2001). Proc. Natl. Acad. Sci. USA, 98, 7783–7788.

  • Chong H and Guan KL . (2003). J. Biol. Chem., 278, 36269–36276.

  • Chong H, Vikis HG and Guan KL . (2003). Cell. Signal., 15, 463–469.

  • Cleveland JL, Troppmair J, Packham G, Askew DS, Lloyd P, Gonzalez-Garcia M, Nunez G, Ihle JN and Rapp UR . (1994). Oncogene, 9, 2217–2226.

  • Cornelis S, Bruynooghe Y, Denecker G, Van Huffel S, Tinton S and Beyaert R . (2000). Mol. Cell, 5, 597–605.

  • Costantini P, Bruey JM, Castedo M, Metivier D, Loeffler M, Susin SA, Ravagnan L, Zamzami N, Garrido C and Kroemer G . (2002). Cell Death Differ., 9, 82–88.

  • Cutler Jr RE, Stephens RM, Saracino MR and Morrison DK . (1998). Proc. Natl. Acad. Sci. USA, 95, 9214–9219.

  • Datta SR, Dudek H, Tao X, Masters S, Fu H, Gotoh Y and Greenberg ME . (1997). Cell, 91, 231–241.

  • Davis RJ . (1994). Trends Biochem. Sci., 19, 470–473.

  • Denecker G, Vercammen D, Steemans M, Vanden Berghe T, Brouckaert G, Van Loo G, Zhivotovsky B, Fiers W, Grooten J, Declercq W and Vandenabeele P . (2001). Cell Death Differ., 8, 829–840.

  • Earnshaw WC, Martins LM and Kaufmann SH . (1999). Annu. Rev. Biochem., 68, 383–424.

  • Emoto Y, Manome Y, Meinhardt G, Kisaki H, Kharbanda S, Robertson M, Ghayur T, Wong WW, Kamen R and Weichselbaum R . (1995). EMBO J., 14, 6148–6156.

  • Garcia-Calvo M, Peterson EP, Rasper DM, Vaillancourt JP, Zamboni R, Nicholson DW and Thornberry NA . (1999). Cell Death Differ., 6, 362–369.

  • Hapel AJ, Warren HS and Hume DA . (1984). Blood, 64, 786–790.

  • Hase T, Muller U, Riezman H and Schatz G . (1984). EMBO J., 3, 3157–3164.

  • Heidecker G, Huleihel M, Cleveland JL, Kolch W, Beck TW, Lloyd P, Pawson T and Rapp UR . (1990). Mol. Cell. Biol., 10, 2503–2512.

  • Huser M, Luckett J, Chiloeches A, Mercer K, Iwobi M, Giblett S, Sun XM, Brown J, Marais R and Pritchard C . (2001). EMBO J., 20, 1940–1951.

  • Jesenberger V, Procyk KJ, Ruth J, Schreiber M, Theussl HC, Wagner EF and Baccarini M . (2001). J. Exp. Med., 193, 353–364.

  • Krajewski S, Krajewska M, Ellerby LM, Welsh K, Xie Z, Deveraux QL, Salvesen GS, Bredesen DE, Rosenthal RE, Fiskum G and Reed JC . (1999). Proc. Natl. Acad. Sci. USA, 96, 5752–5757.

  • Le Mellay V, Troppmair J, Benz R and Rapp UR . (2002). BMC Cell Biol., 3, 14.

  • Li H, Zhu H, Xu CJ and Yuan J . (1998). Cell, 94, 491–501.

  • Li P, Nijhawan D, Budihardjo I, Srinivasula SM, Ahmad M, Alnemri ES and Wang X . (1997). Cell, 91, 479–489.

  • Luo X, Budihardjo I, Zou H, Slaughter C and Wang X . (1998). Cell, 94, 481–490.

  • Majewski M, Nieborowska-Skorska M, Salomoni P, Slupianek A, Reiss K, Trotta R, Calabretta B and Skorski T . (1999). Cancer Res., 59, 2815–2819.

  • Mikula M, Schreiber M, Husak Z, Kucerova L, Ruth J, Wieser R, Zatloukal K, Beug H, Wagner EF and Baccarini M . (2001). EMBO J., 20, 1952–1962.

  • Muzio M, Chinnaiyan AM, Kischkel FC, O’Rourke K, Shevchenko A, Ni J, Scaffidi C, Bretz JD, Zhang M, Gentz R, Mann M, Krammer PH, Peter ME and Dixit VM . (1996). Cell, 85, 817–827.

  • Palacios R and Steinmetz M . (1985). Cell, 41, 727–734.

  • Peruzzi F, Prisco M, Dews M, Salomoni P, Grassilli E, Romano G, Calabretta B and Baserga R . (1999). Mol. Cell. Biol., 19, 7203–7215.

  • Potokar M, Milisav I, Kreft M, Stenovec M and Zorec R . (2003). FEBS Lett., 544, 153–159.

  • Robinson MJ and Cobb MH . (1997). Curr. Opin. Cell Biol., 9, 180–186.

  • Salomoni P, Wasik MA, Riedel RF, Reiss K, Choi JK, Skorski T and Calabretta B . (1998). J. Exp. Med., 187, 1995–2007.

  • Scaffidi C, Fulda S, Srinivasan A, Friesen C, Li F, Tomaselli KJ, Debatin KM, Krammer PH and Peter ME . (1998). EMBO J., 17, 1675–1687.

  • Stanton Jr VP and Cooper GM . (1987). Mol. Cell. Biol., 7, 1171–1179.

  • Stanton Jr VP, Nichols DW, Laudano AP and Cooper GM . (1989). Mol. Cell. Biol., 9, 639–647.

  • Stennicke HR, Deveraux QL, Humke EW, Reed JC, Dixit VM and Salvesen GS . (1999). J. Biol. Chem., 274, 8359–8362.

  • Susin SA, Lorenzo HK, Zamzami N, Marzo I, Brenner C, Larochette N, Prevost MC, Alzari PM and Kroemer G . (1999). J. Exp. Med., 189, 381–394.

  • Thornberry NA, Rano TA, Peterson EP, Rasper DM, Timkey T, Garcia-Calvo M, Houtzager VM, Nordstrom PA, Roy S, Vaillancourt JP, Chapman KT and Nicholson DW . (1997). J. Biol. Chem., 272, 17907–17911.

  • Tran NH and Frost JA . (2003). J. Biol. Chem., 278, 11221–11226.

  • Troppmair J and Rapp UR . (2003). Biochem. Pharmacol., 66, 1341–1345.

  • van Loo G, Saelens X, Matthijssens F, Schotte P, Beyaert R, Declercq W and Vandenabeele P . (2002). Cell Death Differ., 9, 1207–1211.

  • Vander Heiden MG, Chandel NS, Williamson EK, Schumacker PT and Thompson CB . (1997). Cell, 91, 627–637.

  • Vercammen D, Brouckaert G, Denecker G, Van de Craen M, Declercq W, Fiers W and Vandenabeele P . (1998). J. Exp. Med., 188, 919–930.

  • Wang HG, Miyashita T, Takayama S, Sato T, Torigoe T, Krajewski S, Tanaka S, Hovey III L, Troppmair J, Rapp UR and Reed YC . (1994). Oncogene, 9, 2751–2756.

  • Wang HG, Rapp UR and Reed JC . (1996). Cell, 87, 629–638.

  • Wojnowski L, Stancato LF, Zimmer AM, Hahn H, Beck TW, Larner AC, Rapp UR and Zimmer A . (1998). Mech. Dev., 76, 141–149.

  • Zha J, Harada H, Yang E, Jockel J and Korsmeyer SJ . (1996). Cell, 87, 619–628.

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Acknowledgements

The technical assistance of A Meeuws and W Burm is gratefully acknowledged. This work was supported by the ‘Belgian Federation against Cancer’, the ‘Interuniversity Attraction Poles’ (IUAP-V), and the ‘Fund for Scientific Research-Flanders’ (FWO-Vlaanderen). SC is a postdoctoral research associate with the FWO-Vlaanderen.

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Correspondence to Rudi Beyaert.

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Cornelis, S., Bruynooghe, Y., Van Loo, G. et al. Apoptosis of hematopoietic cells induced by growth factor withdrawal is associated with caspase-9 mediated cleavage of Raf-1. Oncogene 24, 1552–1562 (2005). https://doi.org/10.1038/sj.onc.1208401

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