Background: Eukaryotic initiation factor 2B (eIF2B) initiates and regulates translation initiation in eukaryotes. eIF2B gene mutations cause leukoencephalopathy called vanishing white matter disease (VWM) in humans and slow growth (Slg−) and general control derepression (Gcd−) phenotypes in Saccharomyces cerevisiae. Results: To suppress eIF2B mutations, S. cerevisiae genomic DNA library was constructed in high-copy vector (YEp24) and transformed into eIF2B mutant S. cerevisiae strains. The library was screened for wild-type genes rescuing S. cerevisiae (Slg−) and (Gcd−) phenotypes. A genomic clone, Suppressor-I (Sup-I), rescued S. cerevisiae Slg− and Gcd− phenotypes (gcd7-201 gcn2∆). The YEp24/Sup-I construct contained truncated TAN1, full length EMC4, full length YGL230C, and truncated SAP4 genes. Full length EMC4 (chaperone protein) gene was sub-cloned into pEG (KG) yeast expression vector and overexpressed in gcd7-201 gcn2∆ strain which suppressed the Slg− and Gcd− phenotype. A GST-Emc4 fusion protein of 47 kDa was detected by western blotting using α-GST antibodies. Suppression was specific to gcd7-201 gcn2∆ mutation in eIF2Bβ and Gcd1-502 gcn2∆ in eIF2Bγ subunit. Emc4p overexpression also protected the wild type and mutant (gcd7-201 gcn2∆, GCD7 gcn2∆, and GCD7 GCN2∆) strains from H2O2, ethanol, and caffeine stress. Conclusions: Our results suggest that Emc4p is involved in eIF2B-mediated translational regulation under stress and could provide an amenable tool to understand the eIF2B-mediated defects.
- General Control Derepressed (GCD)
- General Control Nonderepressible (GCN)
- S. cerevisiae
PubMed: MeSH publication types
- Journal Article