The nucleotide sequences of the 5'-terminal T1 oligonucleotides of Semliki-Forest-virus 42-S and 26-S RNAs are different.

To study the mechanism of the synthesis of Semliki Forest virus (SFV) 26-S RNA, we have isolated the 5'-terminal 'cap'-containing RNase-T1-resistant oligonucleotide (T1 cap) from the genomic 42-S RNA and from the subgenomic 26-S RNA and determined their nucleotide sequences. The T1 caps were purified from 32P-labelled RNAs on two successive two-dimensional fractionation systems: (a) electrophoresis on cellulose acetate paper followed by homochromatography and (b) two-dimensional polyacrylamide gel electrophoresis. The T1 caps derived from the two RNAs had different mobilities in both systems. Their nucleotide sequence was found to be: m7G(5')ppp-(5')ApUpGp- for the 42-S RNA and m7G(5')ppp(5')ApUpUpGp- for the 26-S RNA, respectively. Thus, it appears that the 26-S RNA is not formed by initiation of the RNA polymerase at the 3' end of the negative-strand template followed by 'cleavage and splicing' or as the result of a 'polymerase jump'. Our results, instead, favour the model of internal initiation of the polymerase on the 42-S negative-strand RNA template.