Unfolded Protein Response Suppression in Yeast by Loss of tRNA Modifications.

Alexander Bruch,Roland Klassen,Raffael Schaffrath

doi:10.3390/genes9110516

Abstract

Modifications in the anticodon loop of transfer RNAs (tRNAs) have been shown to ensure optimal codon translation rates and prevent protein homeostasis defects that arise in response to translational pausing. Consequently, several yeast mutants lacking important anticodon loop modifications were shown to accumulate protein aggregates. Here we analyze whether this includes the activation of the unfolded protein response (UPR), which is commonly triggered by protein aggregation within the endoplasmic reticulum (ER). We demonstrate that two different aggregation prone tRNA modification mutants (elp6 ncs2; elp3 deg1) lacking combinations of 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U: elp3; elp6; ncs2) and pseudouridine (Ψ: deg1) reduce, rather than increase, splicing of HAC1 mRNA, an event normally occurring as a precondition of UPR induction. In addition, tunicamycin (TM) induced HAC1 splicing is strongly impaired in the elp3 deg1 mutant. Strikingly, this mutant displays UPR independent resistance against TM, a phenotype we found to be rescued by overexpression of tRNAGln(UUG), the tRNA species usually carrying the mcm5s2U34 and Ψ38 modifications. Our data indicate that proper tRNA anticodon loop modifications promote rather than impair UPR activation and reveal that protein synthesis and homeostasis defects in their absence do not routinely result in UPR induction but may relieve endogenous ER stress.

Highlights

Transfer RNA is extensively modified to fine-tune the efficiency of translation.Various transfer RNAs (tRNAs) modifications are known to contribute to the maintenance of optimal codon translation rates in yeast by preventing ribosomal pausing during the decoding process [1,2].For example, 5-methoxycarbonylmethyl-2-thiouridine is present at the wobble position in tRNAGln (UUG) and tRNALys (UUU) and is required for efficient translation of the cognate A-ending codons by both tRNAs [2,3,4]
Ribosomal pausing is increased in double mutants lacking mcm5 U in tandem with s2 U, which goes along with a severe protein homeostasis defect including the accumulation of protein aggregates [2]
Loss of different critical tRNA modifications in yeast induces accumulation of protein aggregates [2,16,36]. It has remained unknown whether protein aggregation induced by tRNA defects occurs solely in the cytoplasm, or within the endoplasmic reticulum (ER)

Summary

Introduction

Transfer RNA (tRNA) is extensively modified to fine-tune the efficiency of translation.Various tRNA modifications are known to contribute to the maintenance of optimal codon translation rates in yeast by preventing ribosomal pausing during the decoding process [1,2].For example, 5-methoxycarbonylmethyl-2-thiouridine (mcm s2 U) is present at the wobble position in tRNAGln (UUG) and tRNALys (UUU) and is required for efficient translation of the cognate A-ending codons by both tRNAs [2,3,4]. Transfer RNA (tRNA) is extensively modified to fine-tune the efficiency of translation. Various tRNA modifications are known to contribute to the maintenance of optimal codon translation rates in yeast by preventing ribosomal pausing during the decoding process [1,2]. 5-methoxycarbonylmethyl-2-thiouridine (mcm s2 U) is present at the wobble position in tRNAGln (UUG) and tRNALys (UUU) and is required for efficient translation of the cognate A-ending codons by both tRNAs [2,3,4]. Formation of the mcm s2 U modification requires the Elongator complex and regulatory proteins, as well as Trm9/Trm112 methyl-transferase [5,6,7,8,9]. Wobble uridine thiolation and mcm U modification occur partially independent of each other. Ribosomal pausing is increased in double mutants lacking mcm U in tandem with s2 U (ncs elp6), which goes along with a severe protein homeostasis defect including the accumulation of protein aggregates [2]

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