Complete translation of encephalomyocarditis virus RNA and faithful cleavage of virus-specific proteins in a cell-free system from Krebs-2 cells

Abstract

Received 27 December 1977 1. Introduction There are numerous reports describing translation of encephalomyocarditis (EMC) virus RNA in cell- free systems (c.f. [l-l 0] ). The products formed in these systems, however, did not coincide, upon poly- acrylamide gel electrophoresis, with ‘mature’ virus- specific polypeptides found in EMC ,virus-infected cells. The difference between the in vitro and in vivo products seemed to be primarily due to two factors, non-complete translation of the RNA template and inefficient cleavage of precursor polypeptides formed. Indeed, the major products of in vitro synthesis corresponded to high-molecular-weight polypeptides containing amino acid sequences of capsid proteins, which implied that only a 5’-terminal region of EMC virus RNA was predominantly, or exclusively, trans- lated. The reasons for such an incomplete translation were thought to be either premature termination at certain regions of the template or template degrada- tion, or both [5,6,10]. In systems where these problems were partially overcome, the resulting product did contain, as minor components, poly- peptides corresponding to the non-capsid 3’-terminal portion of viral genome but only in a high-molecular- weight uncleaved form [ 1 l] . The present report describes conditions under which the entire genome of EMC virus appears to be translated and proper cleavage of polypeptide pre- cursors takes place in vitro. The combination of these events results in formation of almost all ‘mature’ virus-specific polypeptides. Although the formation