Defects in tRNA processing and nuclear export induce GCN4 translation independently of phosphorylation of the α subunit of eukaryotic translation initiation factor 2

Induction of GCN4 translation in amino acid-starved cells involves the inhibition of initiator tRNA(Met) binding to eukaryotic translation initiation factor 2 (eIF2) in response to eIF2 phosphorylation by protein kinase GCN2, It was shown previously that GCN4 translation could be induced independently of GCN2 by overexpressing a mutant tRNA(AAC)(Val), (tRNA(Val*)) or the RNA component of RNase MRP encoded by NME1, Here we show that overexpression of the tRNA pseudouridine 55 synthase encoded by PUS4 also leads to translational derepression of GCN4 (Gcd(-) phenotype) independently of eIF2 phosphorylation, Surprisingly, the Gcd(-) phenotype of high-copy-number PUS4 (hcPUS4) did not require PUS4 enzymatic activity, and several lines of evidence indicate that PUS4 overexpression did not diminish functional initiator tRNA(Met) levels. The presence of hcPUS4 or hcNME1 led to the accumulation of certain tRNA precursors, and their Gcd(-) phenotypes were reversed by overexpressing the RNA component of RNase P (RPR1), responsible for 5'-end processing of all tRNAs, Consistently, overexpression of a mutant pre-tRNA(Tyr) that cannot be processed by RNase P had a Gcd(-) phenotype, Interestingly, the Gcd- phenotype of hcPUS4 also was reversed by overexpressing LOS1, required for efficient nuclear export of tRNA, and los1 Delta cells have a Gcd- phenotype, Overproduced PUS4 appears to impede 5'-end processing or export of certain tRNAs in the nucleus in a manner remedied by increased expression of RNase P or LOS1, respectively, The mutant tRNA(Val*) showed nuclear accumulation in otherwise wild-type cells, suggesting a defect in export to the cytoplasm, We propose that yeast contains a nuclear surveillance system that perceives defects in processing or export of tRNA and evokes a reduction in translation initiation at the step of initiator tRNA(Met) binding to the ribosome.