A single base change in the intron of a serine tRNA affects the rate of RNase P cleavage in vitro and suppressor activity in vivo in Saccharomyces cerevisiae.
Differences in the processing of dimeric tRNASer-tRNAMet precursors derived from the Schizosaccharomyces pombe sup9 wild-type and opal suppressor genes can be attributed to conformational alterations in the tRNASer anticodon/intron domain. A comparison of the patterns obtained upon transcription of the sup9+ (wild-type) and sup9-e (opal suppressor) genes in a coupled transcription/processing extract from Saccharomyces cerevisiae reveals that the latter exhibits a greatly reduced efficiency of 5'-end maturation and is susceptible to specific endonucleolytic cleavage(s) within the intron. Free energy calculations indicate that these effects coincide with a destabilization of the wild-type anticodon/intron stem and suggest that the predominant sup9-e conformer lacks secondary structure in this region. Evidence in support of this hypothesis was obtained by analyzing the processing of sup9+ and sup9-e precursors carrying the intron base substitution, G37:10, which destroys and restores, respectively, the base-pairing potential of the proposed secondary structure and comparing the strength and temperature sensitivity of sup9-e and sup9-e G37:10 suppression in vivo in S. cerevisiae. The data indicate that the anticodon/intron structure of tRNA precursors can influence the rate of RNase P cleavage in vitro and affect tRNA expression in vivo.
D001483:Base Sequence; D004722:Endoribonucleases / Q000378:metabolism*; D005800:Genes, Fungal; D009690:Nucleic Acid Conformation; D012346:RNA, Transfer, Amino Acyl / Q000235:genetics* / Q000378:metabolism; D043262:Ribonuclease P; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D013045:Species Specificity; D013489:Suppression, Genetic; D014158:Transcription, Genetic
D012346:RNA, Transfer, Amino Acyl; D004722:Endoribonucleases; D043262:Ribonuclease P