Translation is regulated via the 3' untranslated region of alpha-myosin heavy chain mRNA by calcium but not by its localization.
Posttranscriptional regulation plays an important role in alpha-myosin heavy chain (alpha-MyHC) protein synthesis in cardiac muscle cells. In the present study, we test the effects of calcium and mRNA mislocalization on alpha-MyHC translation in order to determine the mechanism(s) contributing to translational block via the 3' untranslated region (3'UTR). Neonatal rat cardiac myocytes were treated for 6 h with L-isoproterenol (10 microM) to enhance beating, with 10 microM verapamil to block beating and mislocalize mRNA, or with 3 microM colchicine to enhance beating but mislocalize mRNA by depolymerization of the microtubules. In order to determine whether translation is regulated by the 3'UTR, either a control (SV40 3'UTR) or the experimental (alpha-MyHC 3'UTR) was placed after a luciferase reporter gene and transfected into the myocytes. The amount of luciferase protein only decreased significantly in verapamil arrested cells transfected with the alpha-MyHC 3'UTR construct (P < 0.01). To control for the possibility that pharmacological treatments might affect transcription or message stability, we analyzed neomycin and luciferase mRNA levels transcribed from the same transfected plasmid. No significant changes were found with an RNase protection assay. These results suggest that calcium but not mRNA localization regulates protein synthesis and further, this is mediated by the 3' untranslated region of alpha-MyHC.
Nikcevic
G
G
Department of Physiology and Biophysics, University of Illinois at Chicago, 60612-7342, USA.
Perhonen
M
M
Boateng
S Y
SY
Russell
B
B
D020413:3' Untranslated Regions / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D000831:Animals, Newborn; D002118:Calcium / Q000172:deficiency*; D006321:Heart / Q000187:drug effects / Q000254:growth & development*; D018485:Muscle Fibers, Skeletal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000096:biosynthesis* / Q000235:genetics*; D014176:Protein Biosynthesis / Q000502:physiology*; D017434:Protein Structure, Tertiary / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
D020413:3' Untranslated Regions; D012333:RNA, Messenger; D018995:Myosin Heavy Chains; D002118:Calcium