5′‐Terminal 7‐Methylguanosine and mRNA Function

1 Decapped tobacco mosaic virus (TMV) RNA and rabbit globin mRNA were prepared by enzymic treatment of RNAs with nucleotide pyrophosphatase purified from potato. The extent of removal of 5′-terminal 7-methylguanosine 5′-monophosphate (m7GMP) from TMV RNA was at least 97% as estimated by labeling of the 5′ termini in vitro with S-adenosyl[methyl-3H]methionine catalsed by vaccinia virus methyltransferases. 2 The effect of enzymic decapping was compared with the effect of cap analogs on mRNAs translation in a nuclease-treated rabit reticulocyte lysate and in a wheat germ extract. When translation was studied at low K+ concentration, little or no dependence on 5′-terminal 7-methylguanosine was found with either cell-free system. The importance of the 5′-terminal cap for the efficient translation of TMV RNA and globin mRNA increased as the concentration of K+ in a proteinsynthesis system was raised. In a reticulocyte lysate analogs and enzymic decapping had a similar effect on translation. In a wheat germ extract, mRNA decapping resulted in a more pronounced decrease of mRNA activity, presumably due to the increased susceptibility of decapped mRNAs to the nucleases present in this protein synthesis system. 3 The requirement for a 5′-terminal cap was similar for the synthesis of 130 000-Mr and 165 000-Mr polypeptides coded by TMV RNA. This indicates that both proteins may be initiated at the common site close to the 5′ terminus.