The methylation of the 5' terminal
guanosine residue of the cap structure of Semliki Forest virus (SFV) mRNAs has been shown to occur in vitro concomitantly with their synthesis (R. K. Cross and P. J. Gomatos, Virology, 114, 542-554, 1981). The
enzyme responsible for this methylation, a guanine-7-methyltransferase, is associated with the SFV replication complex which contains both the virus-specified polymerase and
RNA template in a mitochondrial pellet fraction, P-15, from infected cell lysates. In the present report, evidence has been obtained demonstrating that a virus-specified function is required for this methylating activity. First, the
methyltransferase enzyme in these infected P-15 extracts has been found to differ in substrate specificity from that of the BHK host cell
enzyme. This
enzyme was able to catalyze the methylation of
GTP to m7GTP in vitro whereas the cellular
enzyme could not methylate
GTP. The incorporation of a methyl group onto
GTP occurred linearly for at least 2 hr at 30 degrees under conditions of neutral pH and added
GTP substrate. Second, a study of the kinetics of appearance of this activity, has demonstrated that the capacity to methylate
GTP did not appear until 1 hr after
infection and reached maximal levels by about 3 hr. Third, de novo
protein synthesis was required. Addition of the
protein synthesis inhibitor,
cycloheximide, prevented the appearance and subsequent increase in the methylating activity. However, once formed the
methyltransferase was found to be stable for at least 3 hr. These results suggest that an early viral function, perhaps a nonstructural
polypeptide is required for this novel guanine-7-methyltransferase activity in SFV infected
cell extracts.