The essential Gcd10p-Gcd14p nuclear complex is required for 1-methyladenosine modification and maturation of initiator methionyl-tRNA

Gcd10p and Gcd14p are essential proteins required for the initiation of pro tein synthesis and translational repression of GCN4 mRNA. The phenotypes of gcd10 mutants were suppressed by high-copy-number IMT genes, encoding init iator methionyl tRNA (tRNA(i)(Met)), or LHP1, encoding the yeast homolog of the human La autoantigen. The gcd10-504 mutation led to a reduction in ste ady-state levels of mature tRNA(i)(Met), attributable to increased turnover rather than decreased synthesis of pre-tRNA(i)(Met). Remarkably, the letha lity of a GCD10 deletion was suppressed by high-copy-number IMT4, indicatin g that its role in expression of mature tRNA(i)(Met) is the essential funct ion of Gcd10p. A gcd14-2 mutant also showed reduced amounts of mature tRNA( i)(Met), but in addition, displayed a defect in pre-tRNA(i)(Met) processing . Gcd10p and Gcd14p were found to be subunits of a protein complex with pro minent nuclear localization, suggesting a direct role in tRNA(i)(Met) matur ation. The chromatographic behavior of elongator and initiator tRNA(Met) on a RPC-5 column indicated that both species are altered structurally in gcd 10 Delta cells, and analysis of base modifications revealed that 1-methylad enosine (m(1)A) is undetectable in gcd10 Delta tRNA. Interestingly, gcd10 a nd gcd14 mutations had no effect on processing or accumulation of elongator tRNA(Met), which also contains m(1)A at position 58, suggesting a unique r equirement for this base modification in initiator maturation.