Sequence relationships between the genome and the intracellular RNA species of standard and defective-interfering semliki forest virus

Abstract

Oligonucleotide fingerprinting on slabs of polyacrylamide gel (De Wachter & Fiers, 1972) has been used to investigate the sequence relationships between the 42 S RNA genome of standard Semliki Forest virus and the intracellular virus-specified single-stranded RNAs isolated from baby hampster kidney cells infected with standard virus and co-infected with standard virus and defective-interfering particles. For the intracellular RNAs of standard virus-infected cells these studies show that the nucleotide sequence of the 26 S RNA, which acts as messenger RNA for the structural proteins of the virus particle Clegg and Kennedy, 1975a , Clegg and Kennedy, 1975b , is located at the 3′ end of the 42 S RNA and that the 38 S and 33 S RNAs which appear as minor species on RNA polyacrylamide gels run under non-denaturing conditions, are conformational variants of the 42 S and 26 S RNAs, respectively. Studies on the two RNAs of defective-interfering particles isolated from cells co-infected with standard virus and defective-interfering particles indicate extensive sequence homology between the defective-interfering RNAs, and show that both RNAs contain nucleotide sequences present in both the structural and non-structural genes of the 42 S RNA. Furthermore, by determining the order (5′→3′) of the characteristic oligonucleotides of the 42 S RNA, and by digestion with snake venom exonuclease, it is concluded that the nucleotide sequences of the defective-interfering RNAs are derived from the 5′ and 3′ extremities of the 42 S RNA; about 75% from the 5′ end and 25% from the 3′ end. The significance of this observation in relation to the genesis of defective-interfering particles and to the mechanism of interference by the defective-interfering RNAs is discussed.