Nuclear surveillance and degradation of hypomodified initiator tRNA^Met in S. cerevisiae

Abstract

The tRNA m¹A58 methyltransferase is composed of two subunits encoded by the essential genes TRM6 and TRM61 (formerly GCD10 and GCD14). The trm6-504 mutation results in a defective m¹A methyltransferase (Mtase) and a temperature-sensitive growth phenotype that is attributable to the absence of m¹A58 and consequential tRNA_i^Met instability. We used a genetic approach to identify the genes responsible for tRNA_i^Met degradation in trm6 cells. Three recessive extragenic mutations that suppress trm6-504 mutant phenotypes and restore hypomodified tRNA_i^Met to near normal levels were identified. The wild-type allele of one suppressor, DIS3/RRP44, encodes a 3′-5′ exoribonuclease and a member of the multisubunit exosome complex. We provide evidence that a functional nuclear exosome is required for the degradation of tRNA_i^Met lacking m¹A58. A second suppressor gene encodes Trf4p, a DNA polymerase (pol σ) with poly(A) polymerase activity. Whereas deletion of TRF4 leads to stabilization of tRNA_i^Met, overexpression of Trf4p destabilizes the hypomodified tRNA_i^Met in trm6 cells. The hypomodified, but not wild-type, pre-tRNA_i^Met accumulates as a polyadenylated species, whose abundance and length distribution both increase upon Trf4p overexpression. These data indicate that a tRNA surveillance pathway exists in yeast that requires Trf4p and the exosome for polyadenylation and degradation of hypomodified pre-tRNA_i^Met.

Keywords

• Transfer RNA
• exosome
• RNA processing
• RNA turnover
• tRNA modification