Molecular and Cellular Biochemistry

, Volume 275, Issue 1–2, pp 15–24 | Cite as

PKC alpha-dependent regulation of the IGF1 receptor in adult and embryonic rat cardiomyocytes

  • Ruchita Maniar
  • Anna Pecherskaya
  • Richard Ila
  • Michele Solem


In both, the adult rat ventricular cardiomyocytes and the embryonic rat heart cell line, H9c2, acute exposure to IGF1 resulted in activation of the IGF1 receptor’s internal tyrosine kinase, and this was completely blocked by the PKC alpha inhibitor, Gö6976. In addition, RNA interference using siRNA mediated gene silencing of PKC alpha–inhibited IGF1 receptor activity and blocked PKC alpha expression in H9c2 cells. Biochemical experiments demonstrate that PKC alpha is associated with the IGF1R (beta subunit) only after acute IGF1 exposure, and this may suggest that there is a direct interaction and possibly a PKC alpha phosphorylation site within the internal IGF1 receptor domain. The downstream effects of blocking PKC alpha activity by exposure to Gö6976 include inhibition of IGF1-stimuated PI3 kinase activity and reduced IGF1-stimulated c-fos expression in the adult cardiomyocytes. Previously, the laboratory has reported that IGF1 activates PKC alpha in adult rat cardiomyocytes, and that PKC alpha activity is required for IGF1-dependent Erk/Erk2 activity and protein synthesis. Here, it is shown that IGF1-dependent protein synthesis is completely blocked by PD98059, indicating that the Raf-Mek-Erk cascade is required for IGF1’s anabolic activity. Pretreatment with LY294002, a specific inhibitor of PI3 kinase, blocked IGF1-stimulated Erk1/Erk2 activity; therefore, PI3 kinase may also be required for IGF1-dependent protein synthesis. In H9c2 cells, coincubation with PMA lead to an increase in the rate of the IGF1 receptor activation, and this may further implicate a role for PKC in regulating the IGF1R. In conclusion, PKC alpha plays an essential role in the IGF1-signaling cascade, including the regulation of key signaling proteins involved in cell signaling and gene expression, and this may primarily be due to PKC alpha directly regulating the IGF1R.


cardiomyocytes diabetes insulin-like growth factor 1 protein kinase C tyrosine kinase protein synthesis gene transcription 


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  1. 1.
    Buerke M, Murohara T, Skurk S, Nuss C, Tomaselli K, Lefer AM: Cardioprotective effect of insulin-like growth factor 1 in myocardial ischemia followed by reperfusion. Proc Natl Acad Sci USA 92: 8031–8035, 1995Google Scholar
  2. 2.
    Foncea R, Andersson M, Ketterman A, Blakesley V, Sapag-Hagar M, Sugden PH, LeRoith D, Lavandero S: Insulin-like growth factor 1 rapidly activates multiple signal transduction pathways in cultured rat cardiac myocytes. J Biol Chem 272: 19115–19124, 1997Google Scholar
  3. 3.
    Von Lewinski D, Voss K, Hulsmann S, Kogler H, Pieske B: Insulin-like growth factor 1 exerts calcium-dependent positive inotropic effects in failing human myocardium. Circ Res 92: 169–176, 2003Google Scholar
  4. 4.
    Solem ML, Thomas AP: Modulation of the cardiac Ca2+ channel by IGF1. Biochem Biophys Res Commun 252: 151–155, 1998Google Scholar
  5. 5.
    Hubbard SR: Structural analysis of receptor tyrosine kinases. Prog Biophys Mol Biol 71: 343–358, 1999Google Scholar
  6. 6.
    Strack V, Hennige AM, Kruzfeldt J, Bossenmaier B, Klein H, Keller M, Lammers R, Haring HU: Serine residues 994 and 1023/25 are important for insulin receptor kinase inhibition by protein kinase C isoforms beta2 and theta. Diabetologia 43: 443–449, 2000Google Scholar
  7. 7.
    Strack V, Krutzfeldt J, Kellerer M, Ullrich A, Lammers R, Haring HU: The protein tyrosine phosphatase SHP2 is phosphorylated on serine residues 576 and 591 by protein kinase C isoforms alpha, beta1, beta2 and eta. Biochemistry 41: 603–608, 2002Google Scholar
  8. 8.
    Ping P, Zhang J, Zheng YT, Li RCX, Dawn D, Tan XL, Takano H, Balafanova P, Bolli R: Demonstration of selective protein kinase C-dependent activation of Src and Lck tyrosine kinases during ischemic preconditioning in conscious rabbits. Circ Res 85: 542–550, 1999Google Scholar
  9. 9.
    Ravichandran LV, Esposito DI, Chen J, Quon MJ: Protein kinase C zeta phosphorylates insulin receptor substrate-1 and impairs its ability to activate phosphatidylinositol 3-kinase in response to insulin. J Biol Chem 276: 3543–3549, 2001Google Scholar
  10. 10.
    Aguirre V, Werner ED, Giraud J, Lee YL, Shoelson S, White M: Phosphorylation of ser307 in the insulin receptor substrate-1 blocks interactions with the insulin receptor and inhibits insulin action. J Biol Chem 277: 1531–1537, 2002Google Scholar
  11. 11.
    Ho PD, Zechner DK, He H, Dillmann WH, Glembotski CC, McDonough PM: The Raf-MEK-ERK cascade represents a common pathway for alteration of intracellular calcium by ras and protein kinase C in cardiac myocytes. J Biol Chem 273: 21730–2173, 1999Google Scholar
  12. 12.
    Aikawa R, Nawano M, Gu Y, Katagiri H, Asano T, Zhu W, Nagai R, Komuro I: Insulin prevents cardiomyocytes from oxidative stress-induced apoptosis through activation of PI3 kinase/Akt. Circulation 102: 2873–2879, 2000Google Scholar
  13. 13.
    Condorelli G, Drusco A, Stassi G, Bellacosa A, Roncarat R, Iaccarino G, Russo MA, Gu Y, Dalton N, Chung C, Latronico MV, Napoli C, Sadoshim J, Croce CM, Ross J: Akt induces enhanced myocardial contactility and cell size in vivo in transgenic mice. Proc Natl Acad Sci USA 19: 312333–312338, 2002Google Scholar
  14. 14.
    Pecherskaya P, Solem M: Insulin-like Growth Factor 1 activates PKC alpha-dependent protein synthesis in adult rat cardiomyocytes. Mol Cell Biol Res Commun 42: 741–747, 2000Google Scholar
  15. 15.
    O’Rourke B, Reibel D, Thomas A: High-speed digital imaging of cytosolic Ca2+ and contraction in single cardiomyocytes. Am J Physiol 259: H230–H242, 1990Google Scholar
  16. 16.
    Renard DC, Delaville F, Thomas AP: Inhibitory effects of cocaine on Ca2+ transients and contraction in single cardiomyocytes. Am J Physiol 266: H555–H567, 1999Google Scholar
  17. 17.
    Fuller SJ, Mynett JR, Sugden PH: Stimulation of cardiac protein synthesis by IGF1. Biochem J 1282: 85–90, 1992Google Scholar
  18. 18.
    Letiges M, Plomann M, Standaert ML, Bandyopadhyay G, Sajan MP, Kanoh R, Farese RV: Knockout of PKC alpha enhances insulin signaling through PI3K. Mol Endocrin 216: 847–858, 2002Google Scholar
  19. 19.
    Martiny-Baron G, Kazanietz MG, Mischak H, Blumberg PM, Kochs G, Hug H, Marme D, Schachtele C: Selective inhibition of protein kinase C isozymes by the indolocarbazole Gö6976. J Biol Chem 268: 9194–9197, 1993Google Scholar
  20. 20.
    Shepherd PR, Nave BT, O’Rahilly S: The role of phosphoinositide 3-kinase in insulin signaling. J Mol Endorin 17: 175–184, 1996Google Scholar
  21. 21.
    Li J, DeFea K, Roth RA: Modulation of insulin receptor substrate 1 tyrosine phosphorylation by an Akt/phosphatidylinositol 3-kinse pathway. J Biol Chem 14: 9351–9356, 1999Google Scholar
  22. 22.
    Sykiotis GP, Papvassiliou AG: Serine phosphorylation of insulin receptor substrate 1: A novel target or the reversal of insulin resistance. Mol Endocrin 15: 1864–1869, 2000Google Scholar
  23. 23.
    Hong F, Moon K, Kim SS, Kim YS, Choi YK, Bae YS, Suh PG, Ryu SH, Choi EJ, Ha J, Kim SS: Role of phospholipase C gamma1 in insulin-like growth factor I-induced muscle differentiation of H9c2 cardiac myocblast. Biochem Biophys Res Commun 282: 816–822, 2002Google Scholar
  24. 24.
    Rao GN, Delafontaine P, Runge MS: Thrombin stimulates phosphorylation of insulin-like growth factor-1 receptor, insulin receptor substrate-1, and phospholipase C-gamma 1 in rat aortic smooth muscle cells. J Biol Chem 270: 27871–27875, 1995Google Scholar
  25. 25.
    Freeston NS, Ribaric S, Mason WT: The effect of insulin-like growth factor-1 on adult rat cardiac contractility. Mol Cell Biochem 163: 223–229, 1993Google Scholar
  26. 26.
    Donath MY, Gosteli PY, Hauri C, Froesch ER, Zapf J: Insulin-like growth factor I stimulates myofibrillar genes and modulates atrial natriuetic factor mRNA in rat heart. Eur J Endocrin 137: 309–315, 1997Google Scholar
  27. 27.
    Lui TJ, Lai HC, Weihua C, Wang S, Ping PH: Developing a strategy to define the effects of insulin-like growth factor 1 on gene expression profile in cardiomyocytes. Circ Res 88: 12231–1238, 2001Google Scholar
  28. 28.
    Danielsen AG, Liu F, Hosomi F, Shii K, Roth RA: Activation of PKC alpha inhibits signaling by members of the insulin receptor family. J Biol Chem 270: 21600–21605, 1995Google Scholar
  29. 29.
    Braz JC, Buen OH, De Windt LJ, Molkentin JD: PKC alpha regulates the hypertrophic growth of cardiomyocytes through extracellular signal-regulated kinase1/2 (ERK1/2). J Cell Biol 156: 905–919, 2002Google Scholar
  30. 30.
    Heidenreich KA, Zeppelin T, Robinson LJ: Insulin and insulin-like growth factor I induce c-fos expression in postmitotic neurons by a protein kinase C-dependent pathway. J Biol Chem 268: 14663–14670, 1993Google Scholar
  31. 31.
    Ververis JJ, Ku L, Delafontaine P: Regulation of insulin-like growth factor I receptors in vascular smooth muscle cells by growth factors and phorbol esters. Circ Res 72: 1285–1292, 1993Google Scholar
  32. 32.
    Jacobs S, Sahyoun NE, Saltiel AR, Cuatrecasas P: Phorbol esters stimulate the phosphorylation of receptors for insulin and somatomedin C. Proc Natl Acad Sci USA 80: 6211–6213, 1983Google Scholar
  33. 33.
    Braz JC, Gregory K, Pathak A, Zhao W, Sahin B, Klevitsk R, Kimball T, Lorenz JN, Nairn AC, Liggett ST, Bodi I, Wang S, Schwartz A, Lakatta EG, DePaoli-Roach AA, Robbins J, Hewett T, Bibb J, Westfall M, Kranias E, Molkentin JD: PKC-α regulates cardiac contractility and propensity toward heart failure. Nat Med 10: 248–254, 2004Google Scholar
  34. 34.
    Sundgren NC, Giraud GD, Schultz JM, Lasarev M, Stork P, Thornbur KL: Extracellular signal-regulated kinase and phosphoinositol-3 kinase mediate IGF-1 induced proliferation of fetal sheep cardiomyocytes. Am J Physiol Reg Integrat Comp Physiol 285: R1481–R1489, 2003Google Scholar
  35. 35.
    Rohra DK, Yamakuni T, Furukawa KI, Ishii N, Shinkawa T, Isobe T, Ohizumi Y: Stimulated tyrosine phosphorylation of PI3 kinase causes acidic pH-induced contraction in spontaneously hypertensive rat aorta. JPET 303: 1255–1264, 2002Google Scholar
  36. 36.
    Braiman L, Alt A, Kuroki T, Ohba M, Bak A, Tennenbaum T, Sampson SR: Insulin induces specific interaction between insulin receptor and protein kinase delta in primary cultured skeletal muscle. Mol Endocrin 15: 565–574, 2001Google Scholar
  37. 37.
    el Mabrouk M, Touyz R, Schiffin EL: Differential ANG II–dependent growth activation pathways in mesenteric artery smooth muscle cells from SHR. Am J Physiol Heart Circ Physiol 281: H30–H39, 2001Google Scholar
  38. 38.
    Vlahos CJ, Matter WF, Hui KY, Brown RF: A specific inhibitor of phosphatidylinositol 3-kinase, 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002). J Biol Chem 269: 5241–5248, 1994Google Scholar
  39. 39.
    Wennstrom S, Downward J: Role of phosphoinositide 3 kinase in activation of ras and mitogen-activated protein kinase by epidermal growth factor. Mol Cell Bio 19: 4279–4288, 1999Google Scholar

Copyright information

© Springer Science + Business Media, Inc. 2005

Authors and Affiliations

  • Ruchita Maniar
    • 1
  • Anna Pecherskaya
    • 1
  • Richard Ila
    • 1
  • Michele Solem
    • 1
    • 2
  1. 1.Department of Pathology, Anatomy and Cell BiologyThomas Jefferson UniversityPhiladelphiaU.S.A.
  2. 2.Department of Pathology, Anatomy and Cell BiologyThomas Jefferson UniversityPhiladelphiaU.S.A.

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