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

Induction of GCN4 translation in amino acid-starved cells involves the inhi bition 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 independent ly of GCN2 by overexpressing a mutant tRNA(AAC)(Val), (tRNA(Val*)) or the R NA component of RNase MRP encoded by NME1, Here we show that overexpression of the tRNA pseudouridine 55 synthase encoded by PUS4 also leads to transl ational derepression of GCN4 (Gcd(-) phenotype) independently of eIF2 phosp horylation, Surprisingly, the Gcd(-) phenotype of high-copy-number PUS4 (hc PUS4) did not require PUS4 enzymatic activity, and several lines of evidenc e indicate that PUS4 overexpression did not diminish functional initiator t RNA(Met) levels. The presence of hcPUS4 or hcNME1 led to the accumulation o f certain tRNA precursors, and their Gcd(-) phenotypes were reversed by ove rexpressing the RNA component of RNase P (RPR1), responsible for 5'-end pro cessing 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, re quired for efficient nuclear export of tRNA, and los1 Delta cells have a Gc d- phenotype, Overproduced PUS4 appears to impede 5'-end processing or expo rt of certain tRNAs in the nucleus in a manner remedied by increased expres sion of RNase P or LOS1, respectively, The mutant tRNA(Val*) showed nuclear accumulation in otherwise wild-type cells, suggesting a defect in export t o the cytoplasm, We propose that yeast contains a nuclear surveillance syst em that perceives defects in processing or export of tRNA and evokes a redu ction in translation initiation at the step of initiator tRNA(Met) binding to the ribosome.