BINDING OF ENCEPHALOMYOCARDITIS VIRUS-RNA POLYMERASE TO THE 3'-NONCODING REGION OF THE VIRAL-RNA IS SPECIFIC AND REQUIRES THE 3'-POLY(A) TAIL

An in vitro RNA bandshift assay has been developed to demonstrate the binding of purified recombinant encephalomyocarditis (EMC) virus RNA-d ependent RNA polymerase (3D(pol)) to the 3'-noncoding region (NCR) and 30 nucleotides of the adjacent 3'-terminal poly(A) tail (3'-NCR(A)) o f EMC virus RNA. The binding of 3D(pol) to the 3'-NCR(A) fragment was specific since four other unrelated proteins including an RNA polymera se did not bind, and unlabeled 3'-NCR(A), but not alpha-globin mRNA, t RNA, or pure poly(A), competed with radiolabeled 3'-NCR(A) for binding . Surprisingly, 3D(pol) failed to bind to the 3'-NCR of EMC virus RNA lacking the poly(A) tail. The results together show that EMC virus RNA template specificity depends only on 3D(pol), the 3'-NCR, and the pol y(A) tail. This suggests that 3'-poly(A) is essential for viral RNA te mplate selection by the EMC virus RNA polymerase.