TY - JOUR
T1 - Saccharomyces cerevisiae ER membrane protein complex subunit 4 (EMC4) plays a crucial role in eIF2B-mediated translation regulation and survival under stress conditions
AU - Sharma, Sonum
AU - Sourirajan, Anuradha
AU - Baumler, David J.
AU - Dev, Kamal
N1 - Publisher Copyright:
© 2020, The Author(s).
PY - 2020/12/1
Y1 - 2020/12/1
N2 - 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.
AB - 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.
KW - Emc4p
KW - General Control Derepressed (GCD)
KW - General Control Nonderepressible (GCN)
KW - Identification
KW - S. cerevisiae
KW - Suppression
KW - VWM
KW - eIF2B
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UR - http://www.scopus.com/inward/citedby.url?scp=85085898019&partnerID=8YFLogxK
U2 - 10.1186/s43141-020-00029-7
DO - 10.1186/s43141-020-00029-7
M3 - Article
C2 - 32476094
AN - SCOPUS:85085898019
SN - 1687-157X
VL - 18
JO - Journal of Genetic Engineering and Biotechnology
JF - Journal of Genetic Engineering and Biotechnology
IS - 1
M1 - 15
ER -