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 hea vy chain (alpha -MyHC) protein synthesis in cardiac muscle cells. In the pr esent study, we test the effects of calcium and mRNA mislocalization on alp ha -MyHC translation in order to determine the mechanism(s) contributing to translational block via the 3' untranslated region (3'UTR). Neonatal rat c ardiac myocytes were treated for 6 h with L-isoproterenol (10 muM) to enhan ce beating, with 10 muM verapamil to block beating and mislocalize mRNA, or with 3 muM colchicine to enhance beating but mislocalize mRNA by depolymer ization of the microtubules. In order to determine whether translation is r egulated by the 3'UTR, either a control (SV40 3'UTR) or the experimental (a lpha -MyHC 3'UTR) was placed after a luciferase reporter gene and transfect ed into the myocytes. The amount of luciferase protein only decreased signi ficantly 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 neo mycin and luciferase mRNA levels transcribed from the same transfected plas mid. No significant changes were found with an RNase protection assay. Thes e results suggest that calcium but not mRNA localization regulates protein synthesis and further, this is mediated by the 3' untranslated region of al pha -MyHC.