The aim of this study was to investigate whether Shp2 (Src homology region 2, phosphatase 2) controls focal adhesion kinase (FAK) activity and its trophic actions in cardiomyocytes. We show that low phosphorylation levels of FAK in nonstretched neonatal rat ventricular myocytes (NRVMs) coincided with a relatively high basal association of FAK with Shp2 and Shp2 phosphatase activity. Cyclic stretch (15% above initial length) enhanced FAK phosphorylation at Tyr397 and reduced FAK/Shp2 association and phosphatase activity in anti-Shp2 precipitates. Recombinant Shp2 C-terminal protein tyrosine phosphatase domain (Shp2-PTP) interacted with nonphosphorylated recombinant FAK and dephosphorylated FAK immunoprecipitated from NRVMs. Depletion of Shp2 by specific small interfering RNA increased the phosphorylation of FAK Tyr397, Src Tyr418, AKT Ser473, TSC2 Thr1462, and S6 kinase Thr389 and induced hypertrophy of nonstretched NRVMs. Inhibition of FAK/Src activity by PP2 {4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine} abolished the phosphorylation of AKT, TSC2, and S6 kinase, as well as the hypertrophy of NRVMs induced by Shp2 depletion. Inhibition of mTOR (mammalian target of rapamycin) with rapamycin blunted the hypertrophy in NRVMs depleted of Shp2. NRVMs treated with PP2 or depleted of FAK by specific small interfering RNA were defective in FAK, Src, extracellular signal-regulated kinase, AKT, TSC2, and S6 kinase phosphorylation, as well as in the hypertrophic response to prolonged stretch. The stretch-induced hypertrophy of NRVMs was also prevented by rapamycin. These findings demonstrate that basal Shp2 tyrosine phosphatase activity controls the size of cardiomyocytes by downregulating a pathway that involves FAK/Src and mTOR signaling pathways. © 2008 American Heart Association, Inc.1038813824Heineke, J., Molkentin, J.D., Regulation of cardiac hypertrophy by intra-cellular signalling pathways (2006) Nat Rev Mol Cell Biol, 7, pp. 589-600Eble, D.M., Strait, J.B., Govindarajan, G., Lou, J., Byron, K.L., Samarel, A.M., Endothelin-induced cardiomyocyte hypertrophy: Role for focal adhesion kinase (2000) Am J Physiol, 278, pp. H1695-H1707Taylor, J.M., Rovin, J.D., Parsons, J.T., A role for focal adhesion kinase in phenylephrine-induced hypertrophy of rat ventricular cardiomyocytes (2000) J Biol Chem, 275, pp. 19250-19257Torsoni, A.S., Constancio, S.S., Nadruz Jr.W, Hanks, S.K., Franchini, K.G., Focal adhesion kinase is activated and mediates the early hypertrophic response to stretch in cardiomyocytes (2003) Circ Res, 93, pp. 140-147Torsoni, A.S., Marin, T.M., Velloso, L.A., Franchini, K.G., RhoA/ROCK signaling is critical to FAK activation by cyclic stretch in cardiomyocytes (2005) Am J Physiol, 289, pp. H1488-H1496Nadruz Jr.W, Corat, M.A., Marin, T.M., Guimaraes Pereira, G.A., Franchini, K.G., Focal adhesion kinase mediates MEF2 and c-Jun activation by stretch: Role in the activation of the cardiac hypertrophic genetic program (2005) Cardiovasc Res, 68, pp. 87-97Franchini, K.G., Torsoni, A.S., Soares, P.H., Saad, M.J., Early activation of the multicomponent signaling complex associated with focal adhesion kinase induced by pressure overload in the rat heart (2000) Circ Res, 87, pp. 558-565Pham, C.G., Harpf, A.E., Keller, R.S., Vu, H.T., Shai, S.Y., Loftus, J.C., Ross, R.S., Striated muscle-specific beta(1D)-integrin and FAK are involved in car-diomyocyte hypertrophic response pathway (2000) Am J Physiol, 279, pp. H2916-H2926Peng, X., Kraus, M.S., Wei, H., Shen, T.L., Pariaut, R., Alcaraz, A., Ji, G., Guan, J.L., Inactivation of focal adhesion kinase in cardiomyocytes promotes eccentric cardiac hypertrophy and fibrosis in mice (2006) J Clin Invest, 116, pp. 217-227Dimichele, L.A., Doherty, J.T., Rojas, M., Beggs, H.E., Reichardt, L.F., MacK, C.P., Taylor, J.M., Myocyte-restricted focal adhesion kinase deletion attenuates pressure overload-induced hypertrophy (2006) Circ Res, 99, pp. 636-645Clemente, C.F., Tornatore, T.F., Theizen, T.H., Deckmann, A.C., Pereira, T.C., Lopes-Cendes, I., Souza, J.R., Franchini, K.G., Targeting focal adhesion kinase with small interfering RNA prevents and reverses load-induced cardiac hypertrophy in mice (2007) Circ Res, 101, pp. 1339-1348Dunty, J.M., Schaller, M.D., The N termini of focal adhesion kinase family members regulate substrate phosphorylation, localization, and cell morphology (2002) J Biol Chem, 277, pp. 45644-45654Fonseca, P.M., Inoue, R.Y., Kobarg, C.B., Crosara-Alberto, D.P., Kobarg, J., Franchini, K.G., Targeting to C-terminal myosin heavy chain may explain mechanotransduction involving focal adhesion kinase in cardiomyocytes (2005) Circ Res, 96, pp. 73-81Lietha, D., Cai, X., Ceccarelli, D.F., Li, Y., Schaller, M.D., Eck, M.J., Structural basis for the autoinhibition of focal adhesion kinase (2007) Cell, 129, pp. 1177-1187Calalb, M.B., Polte, T.R., Hanks, S.K., Tyrosine phosphorylation of focal adhesion kinase at sites in the catalytic domain regulates kinase activity: A role for Src family kinases (1995) Mol Cell Biol, 15, pp. 954-963Hanks, S.K., Ryzhova, L., Shin, N.Y., Brabek, J., Focal adhesion kinase signaling activities and their implications in the control of cell survival and motility (2003) Front Biosci, 8, pp. d982-d996Yu, D.H., Qu, C.K., Henegariu, O., Lu, X., Feng, G.S., Protein-tyrosine phos-phatase Shp-2 regulates cell spreading, migration, and focal adhesion (1998) J Biol Chem, 273, pp. 21125-21131Manes, S., Mira, E., Gomez-Mouton, C., Zhao, Z.J., Lacalle, R.A., Martinez, A.C., Concerted activity of tyrosine phosphatase SHP-2 and focal adhesion kinase in regulation of cell motility (1999) Mol Cell Biol, 19, pp. 3125-3135Von Wichert, G., Haimovich, B., Feng, G.S., Sheetz, M.P., Force-dependent integrin-cytoskeleton linkage formation requires downregulation of focal complex dynamics by Shp2 (2003) EMBO J, 22, pp. 5023-5035Rafiq, K., Kolpakov, M.A., Abdelfettah, M., Streblow, D.N., Hassid, A., Dell'italia, L.J., Sabri, A., Role of protein-tyrosine phosphatase SHP2 in focal adhesion kinase down-regulation during neutrophil cathepsin G-induced cardiomyo-cytes anoikis (2006) J Biol Chem, 281, pp. 19781-19792Neel, B.G., Gu, H., Pao, L., The 'Shp'ing news: SH2 domain-containing tyrosine phosphatases in cell signaling (2003) Trends Biochem Sci, 28, pp. 284-293Shao, W., Orlando, R.C., Awayda, M.S., Bisphosphonates stimulate an endogenous nonselective cation channel in Xenopus oocytes: Potential mechanism of action (2005) Am J Physiol Cell Physiol, 289, pp. C248-C256Oh, E.S., Gu, H., Saxton, T.M., Timms, J.F., Hausdorff, S., Frevert, E.U., Kahn, B.B., Thomas, S.M., Regulation of early events in integrin signaling by protein tyrosine phosphatase SHP-2 (1999) Mol Cell Biol, 19, pp. 3205-3215Zhang, S.Q., Yang, W., Kontaridis, M.I., Bivona, T.G., Wen, G., Araki, T., Luo, J., Neel, B.G., Shp2 regulates SRC family kinase activity and Ras/Erk activation by controlling Csk recruitment (2004) Mol Cell, 13, pp. 341-355Heidkamp, M.C., Bayer, A.L., Kalina, J.A., Eble, D.M., Samarel, A.M., GFP-FRNK disrupts focal adhesions and induces anoikis in neonatal rat ventricular myocytes (2002) Circ Res, 90, pp. 1282-1289Xu, F., Zhao, R., Peng, Y., Guerrah, A., Zhao, Z.J., Association of tyrosine phosphatase SHP-2 with F-actin at low cell densities (2001) J Biol Chem, 276, pp. 29479-29484Schoenwaelder, S.M., Petch, L.A., Williamson, D., Shen, R., Feng, G.S., Burridge, K., The protein tyrosine phosphatase Shp-2 regulates RhoA activity (2000) Curr Biol, 10, pp. 1523-1526Cobb, B.S., Schaller, M.D., Leu, T.H., Parsons, J.T., Stable association of pp60src and pp59fyn with the focal adhesion-associated protein tyrosine kinase, pp125FAK (1994) Mol Cell Biol, 14, pp. 147-155Araki, T., Nawa, H., Neel, B.G., Tyrosyl phosphorylation of Shp2 is required for normal ERK activation in response to some, but not all, growth factors (2003) J Biol Chem, 278, pp. 41677-41684Zito, C.I., Qin, H., Blenis, J., Bennett, A.M., SHP-2 regulates cell growth by controlling the mTOR/S6 kinase 1 pathway (2007) J Biol Chem, 282, pp. 6946-6953Manning, B.D., Tee, A.R., Logsdon, M.N., Blenis, J., Cantley, L.C., Identification of the tuberous sclerosis complex-2 tumor suppressor gene product tuberin as a target of the phosphoinositide 3-kinase/akt pathway (2002) Mol Cell, 10, pp. 151-162Gan, B., Yoo, Y., Guan, J.L., Association of focal adhesion kinase with tuberous sclerosis complex 2 in the regulation of s6 kinase activation and cell growth (2006) J Biol Chem, 281, pp. 37321-37329Crackower, M.A., Oudit, G.Y., Kozieradzki, I., Sarao, R., Sun, H., Sasaki, T., Hirsch, E., Penninger, J.M., Regulation of myocardial contractility and cell size by distinct PI3K-PTEN signaling pathways (2002) Cell, 110, pp. 737-749Shioi, T., McMullen, J.R., Kang, P.M., Douglas, P.S., Obata, T., Franke, T.F., Cantley, L.C., Izumo, S., Akt/protein kinase B promotes organ growth in transgenic mice (2002) Mol Cell Biol, 22, pp. 2799-2809Shioi, T., McMullen, J.R., Tarnavski, O., Converso, K., Sherwood, M.C., Manning, W.J., Izumo, S., Rapamycin attenuates load-induced cardiac hypertrophy in mice (2003) Circulation, 107, pp. 1664-1670Condorelli, G., Drusco, A., Stassi, G., Bellacosa, A., Roncarati, R., Iaccarino, G., Russo, M.A., Ross Jr.J, Akt induces enhanced myocardial contractility and cell size in vivo in transgenic mice (2002) Proc Natl Acad Sci U S A, 99, pp. 12333-12338Ma, L., Chen, Z., Erdjument-Bromage, H., Tempst, P., Pandolfi, P.P., Phosphor-ylation and functional inactivation of TSC2 by Erk implications for tuberous sclerosis and cancer pathogenesis (2005) Cell, 121, pp. 179-193Kontaridis, M.I., Yang, W., Bence, K.K., Cullen, D., Wang, B., Bodyak, N., Ke, Q., Neel, B.G., Deletion of Ptpn11 (Shp2) in cardiomyocytes causes dilated cardiomyopathy via effects on the extracellular signal-regulated kinase/mitogen-activated protein kinase and RhoA signaling pathways (2008) Circulation, 117, pp. 1423-1435Woywodt, A., Welzel, J., Haase, H., Duerholz, A., Wiegand, U., Potratz, J., Sheikhzadeh, A., Cardiomyopathic lentiginosis/LEOPARD syndrome presenting as sudden cardiac arrest (1998) Chest, 113, pp. 1415-1417Tartaglia, M., Gelb, B.D., Noonan syndrome and related disorders: Genetics and pathogenesis (2005) Annu Rev Genomics Hum Genet, 6, pp. 45-68Araki, T., Mohi, M.G., Ismat, F.A., Bronson, R.T., Williams, I.R., Kutok, J.L., Yang, W., Neel, B.G., Mouse model of Noonan syndrome reveals cell type-and gene dosage-dependent effects of Ptpn11 mutation (2004) Nat Med, 10, pp. 849-857