Cytoplasmic processing of myosin heavy chain messenger RNA: evidence provided by using a recombinant DNA plasmid

Abstract

A recombinant DNA plasmid, designated pMHC25, has been constructed that contains structural gene sequences for rat skeletal muscle myosin heavy chain (MHC). The identity of the MHC sequence insert in pMHC25 was determined by muscle-tissue specificity, inhibition of MHC protein synthesis in vitro by hybrid-arrested translation, purification of mRNA that directs the synthesis of MHC protein in vitro, and hybridization to a 33S cytoplasmic mRNA found only in differentiated muscle cells. pMHC25-DNA-excess filter hybridizations were used to show that more than 90% of the newly synthesized MHC mRNA that appears in the cytoplasm of differentiated L6E9 myotubes contains a long 3' poly(A) tail. In contrast, 90% of the MHC mRNA that accumulates in the cytoplasm of these same cells during myogenic differentiation lacks this long 3' poly(A) tail. These results suggest the occurrence of a posttranscriptional event in differentiated L6E9 myotubes that involves the cytoplasmic processing of poly(A)+ MHC mRNA to poly(A)- or poly(A)-short MHC mRNA.