Frameshift suppressor mutations affecting the major glycine transfer RNAs of Saccharomyces cerevisiae

Abstract

Mutations have been identified in Saccharomyces cerevisiae glycine tRNA genes that result in suppression of +1 frameshift mutations in glycine codons. Wild-type and suppressor alleles of genes encoding the two major glycine tRNAs, tRNA(GCC) and tRNA(UCC), were examined in this study. The genes were identified by genetic complementation and by hybridization to a yeast genomic library using purified tRNA probes. tRNA(UCC) is encoded by three genes, whereas approximately 15 genes encode tRNA(GCC). The frameshift suppressor genes suf1+, suf4+ and suf6+ were shown to encode the wild-type tRNA(UCC) tRNA. The suf1+ and suf4+ genes were identical in DNA sequence, whereas the suf6+ gene, whose DNA sequence was not determined, was shown by a hybridization experiment to encode tRNA(UCC). The ultraviolet light-induced SU F1-1 and spontaneous SU F4-1 suppressor mutations were each shown to differ from wild-type at two positions in the anticodon, including a +1 base-pair insertion and a base-pair substitution. These changes resulted in a CCCC four-base anticodon rather than the CCU three-base anticodon found in wild-type. The RNA sequence of tRNA(UCC) was shown to contain a modified uridine in the wobble position. Mutant tRNA(CCCC) isolated from a SU F1-1 strain lacked this modification. Three unlinked genes that encode wild-type tRNA(GCC), suf20+, trn2, and suf17+, were identical in DNA sequence to the previously described suf16+ frameshift suppressor gene. Spontaneous suppressor mutations at the SU F20 and SU F17 loci were analyzed. The SU F20-2 suppressor allele contained a CCCC anticodon. This allele was derived in two serial selections through two independent mutational events, a +1 base insertion and a base substitution in the anticodon. Presumably, the original suppressor allele, SU F20-1, contained the single base insertion. The SU F17-1 suppressor allele also contained a CCCC anticodon resulting from two mutations, a +1 insertion and a base substitution. However, this allele contained an additional base substitution at position 33 adjacent to the 5' side of the four-base anticodon. The possible origin and significance of multiple mutations leading to frameshift suppression is discussed.