tRNA synthesis: identification of in vivo precursor tRNAs from parental and mutant yeast strains

In vivo yeast precursor tRNAs have been identified using a modification of the Northern-hybridization procedure. Two species of pre-$tRNA2Ser$ and 2 species of pre-tRNA$Serminor$ and 2 species of pre-tRNA_minor^Ser have been found in all yeast strains examined, including parental strains and strains harboring mutations affecting tRNA function. One of the tRNA ^1yr precursors and one of the pre-tRNA$SerUCG$ are the same size as the unspliced pre-tRNAs which accumulate in the yeast mutant rnal. The in vivo tRNA^1yr precursors detected in these studies also appear similar with the RNA species identified when cloned yeast tRNA^1yr is transcribed and processed by Xenopus oocytes and/or Xenopus extracts. We have also studied the precursor and mature tRNA^1yr species from 22 mutants which contain mutations in the SUP4 tyrosine-inserting suppressor locus. The RNA from 2 mutants mapping at the G52 position showed an aberrantly migrating “mature” tRNA^1yr. Although several of those cloned mutant genes showed transcript products of altered size in in vitro transcription studies (1), we did not detect such altered transcripts in vivo.