Characterization of the simian virus 40-specific messenger RNAs isolated from HeLa cells infected with the non-defective adenovirus 2-simian virus 40 hybrid viruses Ad2 +ND2 and Ad2 +ND4

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Previous work has shown that cells infected with the non-defective adenovirus 2-simian virus 40 hybrid viruses, Ad2 +ND2 and Ad2 +ND4 synthesize more than one SV40 § § Abbreviations used: SV40, simian virus 40; Ad2, adenovirus 2; Ad2 +ND1, −2, −3, −4 or −5, non-defective adenovirus 2-simian virus 40 hybrid 1, 2, 3, 4 or 5; fmet-tRNA f met, formyl methionyl tRNA f met. large T antigen-related protein. These proteins overlap in amino acid sequence and have their carboxy-terminal sequences in common (Mann et al., 1977). We have characterized the messenger RNAs coding for these SV40-specific proteins. By translating in vitro SV40-specific mRNA isolated from cells infected with these viruses we have shown that each SV40-specific protein can incorporate 35S-labeled formyl methionine at its N-terminus donated by [ 35S]-fmet-tRNA f met, demonstrating that each protein results from a de novo initiation event. Furthermore, analysis of the N-terminal tryptic peptides of these proteins indicates that each protein has a unique N-terminal peptide and therefore a unique initiation site for protein synthesis, with the possible exception of the 74,000 and 95,000 molecular weight proteins, which may have the same N-terminal sequence. Therefore, these proteins cannot be derived by proteolytic cleavage of a large precursor protein. The messenger activities for many of the hybrid virus proteins can be resolved by gel electrophoresis, demonstrating the presence of multiple SV40-specific mRNA species. This result is consistent with the possibility that each SV40-specific protein is coded by a distinct species of RNA.