A NORMAL TRNA(ARG) COMPLEMENTS THE RAD18X MUTATION IN SCHIZOSACCHAROMYCES-POMBE

Abstract

A clone that complements the rad18x mutation of Schizosaccharomyces pombe has been isolated from the screening of yeast genomic DNA library. This clone with a 4.6 kb DNA insert was named pPS1 and appeared to partially complement the UV and methyl methanesulfonate (MMS) sensitivity of rad18 mutatant The region responsible for the complementing activity on rad18x mutation was located within a 1.0 kb BamHI-SmaI fragment within the pPS1. Northern blot and nucleotide sequence analyses of this fragment revealed that the DNA insert of pPS1 carries a normal tRNA gene consisting of 73 base pairs without an intron. Its anticodon sequence is ACG which primarily decodes CGT (arginine). A possible RNA polymerase III termination site is present downstream from the end of the tRNA. The KJ3 strain of S. pombe which has a known amber (UAG) mutation was suppressed by the transformation with pPS1. These results suggest that a normal tRNA(Arg) in 1.0 kb BamHI-SmaI fragment of pPS1 could overcome the nonsense mutation in the rad18 mutant when it is expressed in a sufficient quantity.