In vivo modulation of yeast tRNA gene expression by 5'-flanking sequences.
A pentadecanucleotide sequence, TTTCAACAAATAAGT, contiguous with the 5'-end of Saccharomyces cerevisiae tRNA-Leu3 coding sequence acts as a positive modulator of transcription in a homologous in vitro system. To determine whether modulation also takes place in vivo, the amber suppressor forms of tRNA-Leu3 genes with different 5'-flanking sequences were generated by site-specific mutagenesis and cloned into YCp19, a yeast vector maintained at 1-2 copies per cell. These plasmids were transformed into S. cerevisiae strains marked with amber mutations lys2-801, met8-1, and tyr7-1. The ability of the tRNA-Leu3 amber suppressor genes (tDNA-Leu3A) to suppress functionally lys2-801 and tyr7-1 mutations in the yeast host strain correlated well with template activities measured in vitro. We conclude that the plasmid-borne tRNA gene acts as an effective suppressor from the plasmid and the conserved pentadecanucleotide sequence modulated the expression of yeast tRNA-Leu3 in vivo as well as in vitro. This regulatory sequence if found associated with genes coding for a number of tRNAs which are abundant in yeast. We postulate that this sequence represents a mechanism by which production of specific tRNAs can be enhanced to match demand created by codon use preferences.
D001483:Base Sequence; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics; D005786:Gene Expression Regulation; D005809:Genes, Regulator; D007930:Leucine; D009154:Mutation; D009690:Nucleic Acid Conformation; D012331:RNA, Fungal / Q000235:genetics*; D012343:RNA, Transfer / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D014158:Transcription, Genetic
D004271:DNA, Fungal; D012331:RNA, Fungal; D012343:RNA, Transfer; D007930:Leucine