Sympathetic control of cardiac myosin heavy chain gene expression
Several neuroendocrine factors have been shown to influence the muscle phenotype. Various physiological reports have suggested the role of adrenergic nervous system for cardiac myosin heavy chain (MHC) expression. We have used cultured fetal rat heart myocytes to investigate the role of cAMP on the α- and β-MHC gene expression. In low density cultures, addition of 1 mM 8 Br cAMP resulted in up regulation of α-MHC and down regulation of β-MHC mRNA. This antithetic effect of cAMP depends on the basal expression of both MHC transcripts. In transient transfection analysis employing a series of α-MHC gene promoter/reporter constructs, we identified a 13 bp E-box M-CAT hybrid motif (EM element) which conferred a basal muscle specific and cAMP-inducible expression of the α-MHC gene. Data obtained from the mobility gel-shift analysis indicated that one of the factor(s) binding to the EM element is related to troponin T M-CAT binding factor (TEF-1). To test whether the protein binding to this sequence could be a substrate for cAMP-dependent phosphorylation, the cardiac nuclear proteins were preincubated in a kinase reaction buffer either with a catalytic subunit of PKA (CatPKA) or with cAMP, and binding activity of proteins to the EM element was evaluated by mobility gel shift assay. In a concentration dependent manner, a twofold increase in the intensity of the retarded band was observed. Furthermore, at 100 units of CatPKA, an additional band of faster mobility was observed which was not present either when phosphorylated nuclear extract was incubated with alkaline phosphatase or when ATP was absent in kinase reaction buffer. These results strongly suggest that factor(s) binding to the EM element is a substrate for cAMP dependent phosphorylation.
Key wordscardiac myocytes myosin heavy chain gene cAMP phosphorylation
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- 2.Shimizu N, Camoretti-Mercada B, Jakovcic S, Zak R: RNA transcription in heart muscle. In: H.A. Fozzard, E. Haber, R.B. Jennings, A.M. Katz, H.E. Morgan (eds). The Heart and Cardiovascular System, 2nd ed., Raven Press, Ltd, New York, 1992, pp 1525–1538Google Scholar
- 3.Bugaisky LB, Gupta M, Gupta MP, Zak R: In: Cellular and molecular mechanisms of cardiac hypertrophy, In: H.A. Fozzard, E. Haber, R.B. Jennings, A.M. Katz, H.E. Morgan (eds). The Heart and Cardiovascular System, 2nd edn, Raven Press, New York, 1992, pp 1621–1640Google Scholar
- 15.Kawana M, Ischizuka N, Taira A, Kimata S, Hosoda S: Effects of cardiac sympathetic activity on myosin isozymes of rabbit heart. Circulation 80 (Suppl II): 462, 1989Google Scholar
- 16.Advani SV, Malhotra A, Liang D, Geener DL, Buttrick PM, Scheuer J: Swimming attenuates the shift in myosin isoenzymes in the rat heterotopic cardiac isograft. Circulation 80 (Suppl II):297, 1989Google Scholar
- 23.Higuchi R: Recombinant PCR. In: M.A. Iunis, D.H. Gelfand, J.J. Sninsky, T.J. White (eds). PCR Protocols: A Guide to Methods and Applications, Academic Press, New York, pp 177–183, 1990Google Scholar
- 32.Mar JH, Ordahl CP: M-CAT binding factor, a novel transcription factor governing muscle-specific transcription. Mol Cell Biol 12: 619–630, 1990Google Scholar
- 35.Ishiji T, Lack MJ, Parkkinen S, Anderson RD, Hangen TH, Crippe TP, Xiao JH, Davidson I, Chambon P, Turek LP: Transcription enhancer factor (TEF-1) and its cell-specific co-activator activates human papillomavirus E6 and E7 oncogene transcription in keratinocytes and cervical carcinoma. EMBO J 11: 2271–2281, 1992PubMedGoogle Scholar
- 42.Jacob R: Chronic reactions of myocardium at the myofibrillar level. Reflections on ‘adaptation’ and ‘disease’ based on the biology of long-term cardiac overload. In: Jacob R et al (ed.). Cardiac Adaptation of Hemodynamic Overload, Training and Stress. Steinkopff Verlag, Darmstadt pp 3–24, 1983Google Scholar