Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 2003 Apr 1;371(Pt 1):71–79. doi: 10.1042/BJ20021395

Chronic activation of extracellular-signal-regulated protein kinases by phenylephrine is required to elicit a hypertrophic response in cardiac myocytes.

Anthony J Barron 1, Stephen G Finn 1, Stephen J Fuller 1
PMCID: PMC1223261  PMID: 12513686

Abstract

Extracellular-signal-regulated protein kinases (ERKs) are activated rapidly and transiently in response to phenylephrine (PE) and endothelin-1 (ET-1) in cardiac myocytes, but whether this is linked to the subsequent development of the hypertrophic phenotype remains equivocal. To investigate this, we examined the dependence of the hypertrophic response on the length of exposure to PE in neonatal myocyte cultures. In addition to the initial transient activation of ERKs (maximum at 5-10 min), PE (10 microM) induced a second, more prolonged peak of activity several hours later. The activity of a transfected atrial natriuretic factor-luciferase reporter gene was increased 10- to 24-fold by PE. This response was inhibited by the alpha(1)-antagonist prazosin (100 nM) and by U0126 (10 microM) and PD184352 (1 microM), inhibitors of ERK activation, irrespective of whether these were added before or up to 24 h after the addition of PE. Prazosin had no effect on ET-1 (50 nM)-stimulated atrial natriuretic factor-luciferase activity. Protein synthesis was enhanced by 35+/-6% by PE, and this was blocked by prazosin added 1 h after the addition of PE, but decreased only by half when added 8 h after PE. Similarly, PE (48 h) increased myocyte area by 49% and this was prevented by prazosin added 1 h after PE, but decreased only by half when added at 24 h. These results demonstrate that prolonged exposure to PE is required to elicit alterations in gene expression, protein synthesis and cell size, characteristic of hypertrophied myocytes, and they confirm that the initial peak of ERK activity is insufficient to trigger hypertrophic responses.

Full Text

The Full Text of this article is available as a PDF (255.6 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Andersson M. B., Ketterman A. J., Bogoyevitch M. A. Differential regulation of parallel mitogen-activated protein kinases in cardiac myocytes revealed by phosphatase inhibition. Biochem Biophys Res Commun. 1998 Oct 9;251(1):328–333. doi: 10.1006/bbrc.1998.9476. [DOI] [PubMed] [Google Scholar]
  2. Aoki H., Richmond M., Izumo S., Sadoshima J. Specific role of the extracellular signal-regulated kinase pathway in angiotensin II-induced cardiac hypertrophy in vitro. Biochem J. 2000 Apr 1;347(Pt 1):275–284. [PMC free article] [PubMed] [Google Scholar]
  3. Bogoyevitch M. A., Gillespie-Brown J., Ketterman A. J., Fuller S. J., Ben-Levy R., Ashworth A., Marshall C. J., Sugden P. H. Stimulation of the stress-activated mitogen-activated protein kinase subfamilies in perfused heart. p38/RK mitogen-activated protein kinases and c-Jun N-terminal kinases are activated by ischemia/reperfusion. Circ Res. 1996 Aug;79(2):162–173. doi: 10.1161/01.res.79.2.162. [DOI] [PubMed] [Google Scholar]
  4. Bogoyevitch M. A., Glennon P. E., Andersson M. B., Clerk A., Lazou A., Marshall C. J., Parker P. J., Sugden P. H. Endothelin-1 and fibroblast growth factors stimulate the mitogen-activated protein kinase signaling cascade in cardiac myocytes. The potential role of the cascade in the integration of two signaling pathways leading to myocyte hypertrophy. J Biol Chem. 1994 Jan 14;269(2):1110–1119. [PubMed] [Google Scholar]
  5. Bogoyevitch M. A., Glennon P. E., Sugden P. H. Endothelin-1, phorbol esters and phenylephrine stimulate MAP kinase activities in ventricular cardiomyocytes. FEBS Lett. 1993 Feb 15;317(3):271–275. doi: 10.1016/0014-5793(93)81291-7. [DOI] [PubMed] [Google Scholar]
  6. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  7. Bueno O. F., De Windt L. J., Tymitz K. M., Witt S. A., Kimball T. R., Klevitsky R., Hewett T. E., Jones S. P., Lefer D. J., Peng C. F. The MEK1-ERK1/2 signaling pathway promotes compensated cardiac hypertrophy in transgenic mice. EMBO J. 2000 Dec 1;19(23):6341–6350. doi: 10.1093/emboj/19.23.6341. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cherrington J. M., Mocarski E. S. Human cytomegalovirus ie1 transactivates the alpha promoter-enhancer via an 18-base-pair repeat element. J Virol. 1989 Mar;63(3):1435–1440. doi: 10.1128/jvi.63.3.1435-1440.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chien K. R., Knowlton K. U., Zhu H., Chien S. Regulation of cardiac gene expression during myocardial growth and hypertrophy: molecular studies of an adaptive physiologic response. FASEB J. 1991 Dec;5(15):3037–3046. doi: 10.1096/fasebj.5.15.1835945. [DOI] [PubMed] [Google Scholar]
  10. Choukroun G., Hajjar R., Fry S., del Monte F., Haq S., Guerrero J. L., Picard M., Rosenzweig A., Force T. Regulation of cardiac hypertrophy in vivo by the stress-activated protein kinases/c-Jun NH(2)-terminal kinases. J Clin Invest. 1999 Aug;104(4):391–398. doi: 10.1172/JCI6350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Choukroun G., Hajjar R., Kyriakis J. M., Bonventre J. V., Rosenzweig A., Force T. Role of the stress-activated protein kinases in endothelin-induced cardiomyocyte hypertrophy. J Clin Invest. 1998 Oct 1;102(7):1311–1320. doi: 10.1172/JCI3512. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Clerk A., Gillespie-Brown J., Fuller S. J., Sugden P. H. Stimulation of phosphatidylinositol hydrolysis, protein kinase C translocation, and mitogen-activated protein kinase activity by bradykinin in rat ventricular myocytes: dissociation from the hypertrophic response. Biochem J. 1996 Jul 1;317(Pt 1):109–118. doi: 10.1042/bj3170109. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Clerk A., Sugden P. H. Activation of protein kinase cascades in the heart by hypertrophic G protein-coupled receptor agonists. Am J Cardiol. 1999 Jun 17;83(12A):64H–69H. doi: 10.1016/s0002-9149(99)00261-1. [DOI] [PubMed] [Google Scholar]
  14. Clerk Angela, Kemp Timothy J., Harrison Joanne G., Mullen Anthony J., Barton Paul J. R., Sugden Peter H. Up-regulation of c-jun mRNA in cardiac myocytes requires the extracellular signal-regulated kinase cascade, but c-Jun N-terminal kinases are required for efficient up-regulation of c-Jun protein. Biochem J. 2002 Nov 15;368(Pt 1):101–110. doi: 10.1042/BJ20021083. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Cook S. J., Aziz N., McMahon M. The repertoire of fos and jun proteins expressed during the G1 phase of the cell cycle is determined by the duration of mitogen-activated protein kinase activation. Mol Cell Biol. 1999 Jan;19(1):330–341. doi: 10.1128/mcb.19.1.330. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Cowley S., Paterson H., Kemp P., Marshall C. J. Activation of MAP kinase kinase is necessary and sufficient for PC12 differentiation and for transformation of NIH 3T3 cells. Cell. 1994 Jun 17;77(6):841–852. doi: 10.1016/0092-8674(94)90133-3. [DOI] [PubMed] [Google Scholar]
  17. Finn S. G., Dickens M., Fuller S. J. c-Jun N-terminal kinase-interacting protein 1 inhibits gene expression in response to hypertrophic agonists in neonatal rat ventricular myocytes. Biochem J. 2001 Sep 1;358(Pt 2):489–495. doi: 10.1042/0264-6021:3580489. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Fukuzawa Jun, Nishihira Jun, Hasebe Naoyuki, Haneda Takashi, Osaki Junzo, Saito Tetsuya, Nomura Tomoaki, Fujino Takayuki, Wakamiya Nobutaka, Kikuchi Kenjiro. Contribution of macrophage migration inhibitory factor to extracellular signal-regulated kinase activation by oxidative stress in cardiomyocytes. J Biol Chem. 2002 Apr 26;277(28):24889–24895. doi: 10.1074/jbc.M112054200. [DOI] [PubMed] [Google Scholar]
  19. Fuller S. J., Davies E. L., Gillespie-Brown J., Sun H., Tonks N. K. Mitogen-activated protein kinase phosphatase 1 inhibits the stimulation of gene expression by hypertrophic agonists in cardiac myocytes. Biochem J. 1997 Apr 15;323(Pt 2):313–319. doi: 10.1042/bj3230313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Fuller S. J., Finn S. G., Downward J., Sugden P. H. Stimulation of gene expression in neonatal rat ventricular myocytes by Ras is mediated by Ral guanine nucleotide dissociation stimulator (Ral.GDS) and phosphatidylinositol 3-kinase in addition to Raf. Biochem J. 1998 Oct 15;335(Pt 2):241–246. doi: 10.1042/bj3350241. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Gillespie-Brown J., Fuller S. J., Bogoyevitch M. A., Cowley S., Sugden P. H. The mitogen-activated protein kinase kinase MEK1 stimulates a pattern of gene expression typical of the hypertrophic phenotype in rat ventricular cardiomyocytes. J Biol Chem. 1995 Nov 24;270(47):28092–28096. doi: 10.1074/jbc.270.47.28092. [DOI] [PubMed] [Google Scholar]
  22. Glennon P. E., Kaddoura S., Sale E. M., Sale G. J., Fuller S. J., Sugden P. H. Depletion of mitogen-activated protein kinase using an antisense oligodeoxynucleotide approach downregulates the phenylephrine-induced hypertrophic response in rat cardiac myocytes. Circ Res. 1996 Jun;78(6):954–961. doi: 10.1161/01.res.78.6.954. [DOI] [PubMed] [Google Scholar]
  23. Gurjar M. V., Deleon J., Sharma R. V., Bhalla R. C. Role of reactive oxygen species in IL-1 beta-stimulated sustained ERK activation and MMP-9 induction. Am J Physiol Heart Circ Physiol. 2001 Dec;281(6):H2568–H2574. doi: 10.1152/ajpheart.2001.281.6.H2568. [DOI] [PubMed] [Google Scholar]
  24. Hefti M. A., Harder B. A., Eppenberger H. M., Schaub M. C. Signaling pathways in cardiac myocyte hypertrophy. J Mol Cell Cardiol. 1997 Nov;29(11):2873–2892. doi: 10.1006/jmcc.1997.0523. [DOI] [PubMed] [Google Scholar]
  25. Hipskind R. A., Baccarini M., Nordheim A. Transient activation of RAF-1, MEK, and ERK2 coincides kinetically with ternary complex factor phosphorylation and immediate-early gene promoter activity in vivo. Mol Cell Biol. 1994 Sep;14(9):6219–6231. doi: 10.1128/mcb.14.9.6219. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Iwaki K., Sukhatme V. P., Shubeita H. E., Chien K. R. Alpha- and beta-adrenergic stimulation induces distinct patterns of immediate early gene expression in neonatal rat myocardial cells. fos/jun expression is associated with sarcomere assembly; Egr-1 induction is primarily an alpha 1-mediated response. J Biol Chem. 1990 Aug 15;265(23):13809–13817. [PubMed] [Google Scholar]
  27. Kamakura S., Moriguchi T., Nishida E. Activation of the protein kinase ERK5/BMK1 by receptor tyrosine kinases. Identification and characterization of a signaling pathway to the nucleus. J Biol Chem. 1999 Sep 10;274(37):26563–26571. doi: 10.1074/jbc.274.37.26563. [DOI] [PubMed] [Google Scholar]
  28. Knowlton K. U., Baracchini E., Ross R. S., Harris A. N., Henderson S. A., Evans S. M., Glembotski C. C., Chien K. R. Co-regulation of the atrial natriuretic factor and cardiac myosin light chain-2 genes during alpha-adrenergic stimulation of neonatal rat ventricular cells. Identification of cis sequences within an embryonic and a constitutive contractile protein gene which mediate inducible expression. J Biol Chem. 1991 Apr 25;266(12):7759–7768. [PubMed] [Google Scholar]
  29. Mansour S. J., Matten W. T., Hermann A. S., Candia J. M., Rong S., Fukasawa K., Vande Woude G. F., Ahn N. G. Transformation of mammalian cells by constitutively active MAP kinase kinase. Science. 1994 Aug 12;265(5174):966–970. doi: 10.1126/science.8052857. [DOI] [PubMed] [Google Scholar]
  30. Marshall C. J. Specificity of receptor tyrosine kinase signaling: transient versus sustained extracellular signal-regulated kinase activation. Cell. 1995 Jan 27;80(2):179–185. doi: 10.1016/0092-8674(95)90401-8. [DOI] [PubMed] [Google Scholar]
  31. Meloche S., Seuwen K., Pagès G., Pouysségur J. Biphasic and synergistic activation of p44mapk (ERK1) by growth factors: correlation between late phase activation and mitogenicity. Mol Endocrinol. 1992 May;6(5):845–854. doi: 10.1210/mend.6.5.1603090. [DOI] [PubMed] [Google Scholar]
  32. Mody N., Leitch J., Armstrong C., Dixon J., Cohen P. Effects of MAP kinase cascade inhibitors on the MKK5/ERK5 pathway. FEBS Lett. 2001 Jul 27;502(1-2):21–24. doi: 10.1016/s0014-5793(01)02651-5. [DOI] [PubMed] [Google Scholar]
  33. Nguyen T. T., Scimeca J. C., Filloux C., Peraldi P., Carpentier J. L., Van Obberghen E. Co-regulation of the mitogen-activated protein kinase, extracellular signal-regulated kinase 1, and the 90-kDa ribosomal S6 kinase in PC12 cells. Distinct effects of the neurotrophic factor, nerve growth factor, and the mitogenic factor, epidermal growth factor. J Biol Chem. 1993 May 5;268(13):9803–9810. [PubMed] [Google Scholar]
  34. Nicol R. L., Frey N., Pearson G., Cobb M., Richardson J., Olson E. N. Activated MEK5 induces serial assembly of sarcomeres and eccentric cardiac hypertrophy. EMBO J. 2001 Jun 1;20(11):2757–2767. doi: 10.1093/emboj/20.11.2757. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Post G. R., Goldstein D., Thuerauf D. J., Glembotski C. C., Brown J. H. Dissociation of p44 and p42 mitogen-activated protein kinase activation from receptor-induced hypertrophy in neonatal rat ventricular myocytes. J Biol Chem. 1996 Apr 5;271(14):8452–8457. doi: 10.1074/jbc.271.14.8452. [DOI] [PubMed] [Google Scholar]
  36. Ramirez M. T., Sah V. P., Zhao X. L., Hunter J. J., Chien K. R., Brown J. H. The MEKK-JNK pathway is stimulated by alpha1-adrenergic receptor and ras activation and is associated with in vitro and in vivo cardiac hypertrophy. J Biol Chem. 1997 May 30;272(22):14057–14061. doi: 10.1074/jbc.272.22.14057. [DOI] [PubMed] [Google Scholar]
  37. Schlessinger J., Bar-Sagi D. Activation of Ras and other signaling pathways by receptor tyrosine kinases. Cold Spring Harb Symp Quant Biol. 1994;59:173–179. doi: 10.1101/sqb.1994.059.01.021. [DOI] [PubMed] [Google Scholar]
  38. Sugden P. H. Signaling in myocardial hypertrophy: life after calcineurin? Circ Res. 1999 Apr 2;84(6):633–646. doi: 10.1161/01.res.84.6.633. [DOI] [PubMed] [Google Scholar]
  39. Thorburn J., McMahon M., Thorburn A. Raf-1 kinase activity is necessary and sufficient for gene expression changes but not sufficient for cellular morphology changes associated with cardiac myocyte hypertrophy. J Biol Chem. 1994 Dec 2;269(48):30580–30586. [PubMed] [Google Scholar]
  40. Traverse S., Gomez N., Paterson H., Marshall C., Cohen P. Sustained activation of the mitogen-activated protein (MAP) kinase cascade may be required for differentiation of PC12 cells. Comparison of the effects of nerve growth factor and epidermal growth factor. Biochem J. 1992 Dec 1;288(Pt 2):351–355. doi: 10.1042/bj2880351. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Traverse S., Seedorf K., Paterson H., Marshall C. J., Cohen P., Ullrich A. EGF triggers neuronal differentiation of PC12 cells that overexpress the EGF receptor. Curr Biol. 1994 Aug 1;4(8):694–701. doi: 10.1016/s0960-9822(00)00154-8. [DOI] [PubMed] [Google Scholar]
  42. Ueyama T., Kawashima S., Sakoda T., Rikitake Y., Ishida T., Kawai M., Yamashita T., Ishido S., Hotta H., Yokoyama M. Requirement of activation of the extracellular signal-regulated kinase cascade in myocardial cell hypertrophy. J Mol Cell Cardiol. 2000 Jun;32(6):947–960. doi: 10.1006/jmcc.2000.1135. [DOI] [PubMed] [Google Scholar]
  43. Wang Y., Su B., Sah V. P., Brown J. H., Han J., Chien K. R. Cardiac hypertrophy induced by mitogen-activated protein kinase kinase 7, a specific activator for c-Jun NH2-terminal kinase in ventricular muscle cells. J Biol Chem. 1998 Mar 6;273(10):5423–5426. doi: 10.1074/jbc.273.10.5423. [DOI] [PubMed] [Google Scholar]
  44. Yue T. L., Gu J. L., Wang C., Reith A. D., Lee J. C., Mirabile R. C., Kreutz R., Wang Y., Maleeff B., Parsons A. A. Extracellular signal-regulated kinase plays an essential role in hypertrophic agonists, endothelin-1 and phenylephrine-induced cardiomyocyte hypertrophy. J Biol Chem. 2000 Dec 1;275(48):37895–37901. doi: 10.1074/jbc.M007037200. [DOI] [PubMed] [Google Scholar]
  45. Zechner D., Thuerauf D. J., Hanford D. S., McDonough P. M., Glembotski C. C. A role for the p38 mitogen-activated protein kinase pathway in myocardial cell growth, sarcomeric organization, and cardiac-specific gene expression. J Cell Biol. 1997 Oct 6;139(1):115–127. doi: 10.1083/jcb.139.1.115. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES