A missense mutation in the nuclear gene coding for the mitochondrial aspartyl-tRNA synthetase suppresses a mitochondrial tRNA(Asp) mutation.
The nuclear suppressor allele NSM3 in strain FF1210-6C/170-E22 (E22), which suppresses a mutation of the yeast mitochondrial tRNA(Asp)gene in Saccharomyces cerevisiae, was cloned and identified. To isolate the NSM3 allele, a genomic DNA library using the vector YEp13 was constructed from strain E22. Nine YEp13 recombinant plasmids were isolated and shown to suppress the mutation in the mitochondrial tRNA(Asp)gene. These nine plasmids carry a common 4. 5-kb chromosomal DNA fragment which contains an open reading frame coding for yeast mitochondrial aspartyl-tRNA synthetase (AspRS) on the basis of its sequence identity to the MSD1 gene. The comparison of NSM3 DNA sequences between the suppressor and the wild-type version, cloned from the parental strain FF1210-6C/170, revealed a G to A transition that causes the replacement of amino acid serine (AGU) by an asparagine (AAU) at position 388. In experiments switching restriction fragments between the wild type and suppressor versions of the NSM3 gene, the rescue of respiratory deficiency was demonstrated only when the substitution was present in the construct. We conclude that the base substitution causes the respiratory rescue and discuss the possible mechanism as one which enhances interaction between the mutated tRNA(Asp)and the suppressor version of AspRS.
Chiang
C S
CS
Department of Medical Research, Mackay Memorial Hospital, Tamshui, Taipei 251, Taiwan, Republic of China.
Liaw
G J
GJ
D000483:Alleles; D000595:Amino Acid Sequence; D001226:Aspartate-tRNA Ligase / Q000235:genetics*; D002467:Cell Nucleus / Q000235:genetics; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics; D004272:DNA, Mitochondrial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005784:Gene Amplification; D005800:Genes, Fungal; D008928:Mitochondria / Q000201:enzymology; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D010957:Plasmids / Q000235:genetics; D012349:RNA, Transfer, Asp / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics*; D017386:Sequence Homology, Amino Acid; D013489:Suppression, Genetic
D004271:DNA, Fungal; D004272:DNA, Mitochondrial; D012349:RNA, Transfer, Asp; D001226:Aspartate-tRNA Ligase