Autophagy is a highly regulated intracellular process involved in the turnover of most cellular constituents and in the maintenance of cellular homeostasis. In this study, we show that the activity of autophagy increases in H 2O 2 or RasV12-induced senescent fibroblasts. Inhibiting autophagy promotes cell apoptosis in senescent cells, suggesting that autophagy activation plays a cytoprotective role. Furthermore, our data indicate that the increase of autophagy in senescent cells is linked to the activation of transcription factor FoxO3A, which blocks ATP generation by transcriptionally up-regulating the expression of PDK4, an inhibitor of pyruvate dehydrogenase complex, thus leading to AMPK activation and mTOR inhibition. These findings suggest a novel mechanism by which FoxO3A factors can activate autophagy via metabolic alteration.
References
[1]
Serrano, M.; Blasco, M.A. Putting the stress on senescence. Curr. Opin. Cell Biol 2001, 13, 748–753.
[2]
Roninson, I.B. Tumor cell senescence in cancer treatment. Cancer Res 2003, 63, 2705–2715.
[3]
Alexander, K.; Hinds, P.W. Requirement for p27KIP1 in retinoblastoma protein-mediated senescence. Mol. Cell. Biol 2001, 21, 3616–3631.
[4]
Sager, R. Senescence as a mode of tumor suppression. Environ. Health Persp 1991, 93, 59–62.
[5]
Campisi, J. Cellular senescence as a tumor-suppressor mechanism. Trends Cell Biol 2001, 11, S27–S31.
[6]
Donehower, L.A.; Harvey, M.; Slagle, B.L.; Mcarthur, M.J.; Montgomery, C.A.; Butel, J.S.; Bradley, A. Mice deficient for P53 are developmentally normal but susceptible to spontaneous tumors. Nature 1992, 356, 215–221.
Herman-Antosiewicz, A.; Johnson, D.E.; Singh, S.V. Sulforaphane causes autophagy to inhibit release of cytochrome c and apoptosis in human prostate cancer cells. Cancer Res 2006, 66, 5828–5835.
[11]
Lu, X.F.; Jiang, X.G.; Lu, Y.B.; Bai, J.H.; Mao, Z.B. Characterization of a novel positive transcription regulatory element that differentially regulates the insulin-like growth factor binding protein-3 (IGFBP-3) gene in senescent cells. J. Biol. Chem 2005, 280, 22606–22615.
[12]
Bursch, W.; Ellinger, A.; Kienzl, H.; Torok, L.; Pandey, S.; Sikorska, M.; Walker, R.; Hermann, R.S. Active cell death induced by the anti-estrogens tamoxifen and ICI 164 384 in human mammary carcinoma cells (MCF-7) in culture: The role of autophagy. Carcinogenesis 1996, 17, 1595–1607.
[13]
Shimizu, S.; Kanaseki, T.; Mizushima, N.; Mizuta, T.; Arakawa-Kobayashi, S.; Thompson, C.B.; Tsujimoto, Y. Role of Bcl-2 family proteins in a non-apoptotic programmed cell death dependent on autophagy genes. Nat. Cell Biol 2004, 6, 1221–1228.
[14]
Stroikin, Y.; Dalen, H.; Brunk, U.T.; Terman, A. Testing the “garbage” accumulation theory of ageing: Mitotic activity protects cells from death induced by inhibition of autophagy. Biogerontology 2005, 6, 39–47.
[15]
Gerland, L.M.; Peyrol, S.; Lallemand, C.; Branche, R.; Magaud, J.P.; Ffrench, M. Association of increased autophagic inclusions labeled for beta-galactosidase with fibroblastic aging. Exp. Gerontol 2003, 38, 887–895.
[16]
Biederbick, A.; Kern, H.F.; Elsasser, H.P. Monodansylcadaverine (MDC) is a specific in vivo marker for autophagic vacuoles. Eur. J. Cell Biol 1995, 66, 3–14.
[17]
Mizushima, N. Methods for monitoring autophagy. Int. J. Biochem. Cell Biol 2004, 36, 2491–2502.
[18]
Petiot, A.; Ogier-Denis, E.; Blommaart, E.F.; Meijer, A.J.; Codogno, P. Distinct classes of phosphatidylinositol 3′-kinases are involved in signaling pathways that control macroautophagy in HT-29 cells. J. Biol. Chem 2000, 275, 992–998.
[19]
Kihara, A.; Kabeya, Y.; Ohsumi, Y.; Yoshimori, T. Beclin-phosphatidylinositol 3-kinase complex functions at the trans-Golgi network. EMBO Rep 2001, 2, 330–335.
[20]
Tassa, A.; Roux, M.P.; Attaix, D.; Bechet, D.M. Class III phosphoinositide 3-kinase—Beclin1 complex mediates the amino acid-dependent regulation of autophagy in C2C12 myotubes. Biochem. J 2003, 376, 577–586.
[21]
Liang, X.H.; Yu, J.; Brown, K.; Levine, B. Beclin 1 contains a leucine-rich nuclear export signal that is required for its autophagy and tumor suppressor function. Cancer Res 2001, 61, 3443–3449.
[22]
Ravikumar, B.; Vacher, C.; Berger, Z.; Davies, J.E.; Luo, S.; Oroz, L.G.; Scaravilli, F.; Easton, D.F.; Duden, R.; O’Kane, C.J.; Rubinsztein, D.C. Inhibition of mTOR induces autophagy and reduces toxicity of polyglutamine expansions in fly and mouse models of Huntington disease. Nat. Genet 2004, 36, 585–595.
[23]
Inoki, K.; Zhu, T.; Guan, K.L. TSC2 mediates cellular energy response to control cell growth and survival. Cell 2003, 115, 577–590.
[24]
Brugarolas, J.; Lei, K.; Hurley, R.L.; Manning, B.D.; Reiling, J.H.; Hafen, E.; Witters, L.A.; Ellisen, L.W.; Kaelin, W.G., Jr. Regulation of mTOR function in response to hypoxia by REDD1 and the TSC1/TSC2 tumor suppressor complex. Genes Dev 2004, 18, 2893–2904.
[25]
Pullen, N.; Thomas, G. The modular phosphorylation and activation of p70s6k. FEBS Lett 1997, 410, 78–82.
[26]
Arsham, A.M.; Neufeld, T.P. Thinking globally and acting locally with TOR. Curr. Opin. Cell Biol 2006, 18, 589–597.
[27]
Hardie, D.G. The AMP-activated protein kinase pathway—new players upstream and downstream. J. Cell Sci 2004, 117, 5479–5487.
[28]
Alessi, D.R.; Andjelkovic, M.; Caudwell, B.; Cron, P.; Morrice, N.; Cohen, P.; Hemmings, B.A. Mechanism of activation of protein kinase B by insulin and IGF-1. EMBO J 1996, 15, 6541–6551.
[29]
Shaw, R.J.; Kosmatka, M.; Bardeesy, N.; Hurley, R.L.; Witters, L.A.; DePinho, R.A.; Cantley, L.C. The tumor suppressor LKB1 kinase directly activates AMP-activated kinase and regulates apoptosis in response to energy stress. Proc. Natl. Acad. Sci. USA 2004, 101, 3329–3335.
[30]
Shaw, R.J.; Lamia, K.A.; Vasquez, D.; Koo, S.H.; Bardeesy, N.; Depinho, R.A.; Montminy, M.; Cantley, L.C. The kinase LKB1 mediates glucose homeostasis in liver and therapeutic effects of metformin. Science 2005, 310, 1642–1646.
[31]
Sugden, M.C.; Holness, M.J. Mechanisms underlying regulation of the expression and activities of the mammalian pyruvate dehydrogenase kinases. Arch. Physiol. Biochem 2006, 112, 139–149.
[32]
Kwon, H.S.; Huang, B.; Unterman, T.G.; Harris, R.A. Protein kinase B-alpha inhibits human pyruvate dehydrogenase kinase-4 gene induction by dexamethasone through inactivation of FOXO transcription factors. Diabetes 2004, 53, 899–910.
[33]
Jia, K.; Chen, D.; Riddle, D.L. The TOR pathway interacts with the insulin signaling pathway to regulate C. elegans larval development, metabolism and life span. Dev. Biol 2004, 131, 3897–3906.
[34]
Wu, A.L.; Kim, J.H.; Zhang, C.; Unterman, T.G.; Chen, J. Forkhead box protein O1 negatively regulates skeletal myocyte differentiation through degradation of mammalian target of rapamycin pathway components. Endocrinology 2008, 149, 1407–1414.
[35]
Southgate, R.J.; Neill, B.; Prelovsek, O.; El-Osta, A.; Kamei, Y.; Miura, S.; Ezaki, O.; McLoughlin, T.J.; Zhang, W.; Unterman, T.G.; Febbraio, M.A. FOXO1 regulates the expression of 4E-BP1 and inhibits mTOR signaling in mammalian skeletal muscle. J. Biol. Chem 2007, 282, 21176–21186.
[36]
Mammucari, C.; Milan, G.; Romanello, V.; Masiero, E.; Rudolf, R.; Del Piccolo, P.; Burden, S.J.; di Lisi, R.; Sandri, C.; Zhao, J.; Goldberg, A.L.; Schiaffino, S.; Sandri, M. FoxO3 controls autophagy in skeletal muscle in vivo. Cell Metab 2007, 6, 458–471.
[37]
Zhao, J.; Brault, J.J.; Schild, A.; Cao, P.; Sandri, M.; Schiaffino, S.; Lecker, S.H.; Goldberg, A.L. FoxO3 coordinately activates protein degradation by the autophagic/lysosomal and proteasomal pathways in atrophying muscle cells. Cell Metab 2007, 6, 472–483.
[38]
Calnan, D.R.; Brunet, A. The FoxO code. Oncogene 2008, 27, 2276–2288.
[39]
Wang, Y.; Meng, A.; Zhou, D. Inhibition of phosphatidylinostol 3-kinase uncouples H2O2-induced senescent phenotype and cell cycle arrest in normal human diploid fibroblasts. Exp. Cell Res 2004, 298, 188–196.
[40]
Courtois-Cox, S.; Genther Williams, S.M.; Reczek, E.E.; Johnson, B.W.; McGillicuddy, L.T.; Johannessen, C.M.; Hollstein, P.E.; MacCollin, M.; Cichowski, K. A negative feedback signaling network underlies oncogene-induced senescence. Cancer Cell 2006, 10, 459–472.
[41]
Mawal-Dewan, M.; Lorenzini, A.; Frisoni, L.; Zhang, H.; Cristofalo, V.J.; Sell, C. Regulation of collagenase expression during replicative senescence in human fibroblasts by Akt-forkhead signaling. J. Biol. Chem 2002, 277, 7857–7864.
