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- Title
Functions of eIF3 downstream of 48S assembly impact AUG recognition and GCN4 translational control.
- Authors
Nielsen, Klaus H.; Szamecz, Béla; Valášek, Leoš Jivotovskaya, Antonina; Shin, Byung-Sik; Hinnebusch, Alan G.
- Abstract
The binding of eIF2-GTP-tRNAiMet ternary complex (TC) to 40S subunits is impaired in yeast prt1-1 (eIF3b) mutant extracts, but evidence is lacking that TC recruitment is a critical function of eIF3 in vivo. If TC binding was rate-limiting in prt1-1 cells, overexpressing TC should suppress the temperature-sensitive phenotype and GCN4 translation should be strongly derepressed in this mutant, but neither was observed. Rather, GCN4 translation is noninducible in prt1-1 cells, and genetic analysis indicates defective ribosomal scanning between the upstream open reading frames that mediate translational control. prt1-1 cells also show reduced utilization of a near-cognate start codon, implicating eIF3 in AUG selection. Using in vivo cross-linking, we observed accumulation of TC and mRNA/eIF4G on 40S subunits and a 48S ‘halfmer’ in prt1-1 cells. Genetic evidence suggests that 40S-60S subunit joining is not rate-limiting in the prt1-1 mutant. Thus, eIF3b functions between 48S assembly and subunit joining to influence AUG recognition and reinitiation on GCN4 mRNA. Other mutations that disrupt eIF2-eIF3 contacts in the multifactor complex (MFC) diminished 40S-bound TC, indicating that MFC formation enhances 43S assembly in vivo.
- Subjects
GENETIC translation; MESSENGER RNA; PHENOTYPES; RIBOSOMES; CELLS; GENETIC regulation
- Publication
EMBO Journal, 2004, Vol 23, Issue 5, p1166
- ISSN
0261-4189
- Publication type
Article
- DOI
10.1038/sj.emboj.7600116