| Abstract: | Reovirus mRNA synthesis in vitro by the virion-associated RNA polymerase was only slightly (10 to 15%) diminished in the presence of 2 mM S-adenosylethionine. However, methyl group transfer from S-adenosylmethionine (0.05 mM) to the 5'-terminal cap structure, m7GpppGm in this mRNA was markedly inhibited (80%) under these conditions. Replacement of S-adenosylmethionine by S-adenosylethionine (5 mM) yielded mRNAs containing mainly (70%) 5'-terminal e7GpppGe and e7GpppG, but some of the products were unalkylated (5'-GpppG, ppG). The ethylated mRNAs, but not the unalkylated molecules, bound to wheat germ ribosomes and were translated essentially as well as the corresponding methylated mRNAs in wheat germ extracts and in nuclease-treated rabbit reticulocyte lysates. Protein synthesis directed by ethylated mRNAs in wheat germ extract was 80% decreased by 0.1 mM m7GMP. Under conditions of limited initiation, methylated mRNA bound to wheat germ ribosomes preferentially as compared to ethylated mRNA. The results document for the first time the synthesis of ethylated mRNA and support the hypothesis that N7-alkylation of the 5'-guanosine in caps, rather than methylation itself, is important for the enhancing effect of cap on the initiation of eukaryotic protein synthesis. |
