Myosin heavy chain gene expression in neonatal rat heart cells: effects of [Ca2+]i and contractile activity.
To determine if mechanical signals or alterations in intracellular Ca2+ concentration ([Ca2+]i) affect myosin heavy chain (MHC) gene expression in spontaneously beating, neonatal rat ventricular myocytes, contractile activity was inhibited with verapamil, KCl, or 2,3-butanedione monoxime (BDM), and their acute and chronic effects on myocyte shortening, [Ca2+]i, and MHC gene expression were examined. Despite their differing effects on [Ca2+]i, verapamil, KCl, and BDM all inhibited contractile activity and markedly downregulated beta-MHC mRNA levels to 24 +/- 5, 21 +/- 7, and 6 +/- 2% of contracting cells, respectively. In contrast, these inhibitors of contraction upregulated alpha-MHC mRNA levels to 163 +/- 19, 156 +/- 7, and 198 +/- 20% of contracting cells, respectively. Transient transfection with a rat beta-MHC promoter-luciferase expression plasmid demonstrated that all inhibitors of contraction significantly decreased beta-MHC promoter activity. Paradoxically, contractile arrest also inhibited alpha-MHC promoter activity, suggesting that increased alpha-MHC mRNA levels resulted from posttranscriptional mechanisms. Actinomycin D mRNA stability assays indicated that alpha-MHC mRNA half-life was prolonged in noncontracting cells (33 h) compared with contracting myocytes (14 h). Contraction-dependent alterations in MHC gene expression were not dependent on release of angiotensin II or other growth factors into the culture medium. Thus intrinsic mechanical signals rather than alterations in [Ca2+]i regulate alpha-MHC and beta-MHC gene expression by both transcriptional and posttranscriptional mechanisms.
Qi
M
M
Cardiovascular Institute, Loyola University Chicago Strich School of Medicine, Maywood, Illinois 60153, USA.
Puglisi
J L
JL
Byron
K L
KL
Ojamaa
K
K
Klein
I
I
Bers
D M
DM
Samarel
A M
AM
D000804:Angiotensin II / Q000502:physiology; D000818:Animals; D000831:Animals, Newborn / Q000502:physiology*; D002118:Calcium / Q000378:metabolism; D005786:Gene Expression Regulation; D006207:Half-Life; D006321:Heart / Q000502:physiology*; D007425:Intracellular Membranes / Q000378:metabolism; D009200:Myocardial Contraction; D009206:Myocardium / Q000166:cytology / Q000378:metabolism; D018995:Myosin Heavy Chains / Q000235:genetics*; D009994:Osmolar Concentration; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
D012333:RNA, Messenger; D000804:Angiotensin II; D018995:Myosin Heavy Chains; D002118:Calcium