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Saccharomyces cerevisiae ER membrane protein complex subunit 4 (EMC4) plays a crucial role in eIF2B-mediated translation regulation and survival under stress conditions.

ABSTRACT: 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 ?-GST antibodies. Suppression was specific to gcd7-201 gcn2? mutation in eIF2B? and Gcd1-502 gcn2? in eIF2B? 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.

SUBMITTER: Sharma S

PROVIDER: S-EPMC7261713 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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Saccharomyces cerevisiae ER membrane protein complex subunit 4 (EMC4) plays a crucial role in eIF2B-mediated translation regulation and survival under stress conditions.

Sharma Sonum S Sourirajan Anuradha A Baumler David J DJ Dev Kamal K

Journal, genetic engineering & biotechnology 20200601 1

<h4>Background</h4>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<sup>-</sup>) and general control derepression (Gcd<sup>-</sup>) phenotypes in Saccharomyces cerevisiae.<h4>Results</h4>To suppress eIF2B mutations, S. cerevisiae genomic DNA library was constructed in high-copy vector (YEp24) and transformed into eIF2B muta ...[more]

PMID: 32476094

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Project description:Thioredoxins are small proteins that regulate the cellular redox state, prevent oxidative damage, and play an active role in cell repair. Oxidative stress has proven to be of much relevance in biotechnological processes when the metabolism of Saccharomyces cerevisiae is mainly respiratory. During wine yeast starter production, active dry yeast cytosolic thioredoxin Trx2p is a key player in protecting metabolic enzymes from being oxidized by carbonylation. Less is known about the role of redox control during grape juice fermentation. A mutant strain that lacked both cytosolic thioredoxins, Trx1p and Trx2p, was tested for grape juice fermentation. Its growth and sugar consumption were greatly impaired, which indicates the system's relevance under fermentative conditions. A proteomic analysis indicated that deletion of the genes TRX1 and TRX2 caused a reduction in the ribosomal proteins and factors involved in translation elongation in addition to enzymes for glycolysis and amino acid biosynthesis. A metabolomic analysis of the trx1Δ trx2Δ mutant showed an increase in most proteogenic amino acids, phospholipids, and sphingolipids and higher fatty acid desaturase Ole1p content. Low glycolytic activity was behind the reduced growth and fermentative capacity of the thioredoxin deletion strain. All three hexokinases were downregulated in the mutant strain, but total hexokinase activity remained, probably due to posttranslational regulation. Pyruvate kinase Cdc19p presented an early level of aggregation in the trx1Δ trx2Δ mutant, which may contribute to a diminished hexose metabolism and trigger regulatory mechanisms that could influence the level of glycolytic enzymes.IMPORTANCE Oxidative stress is a common hazardous condition that cells have to face in their lifetime. Oxidative damage may diminish cell vitality and viability by reducing metabolism and eventually leading to aging and ultimate death. Wine yeast Saccharomyces cerevisiae also faces oxidative attack during its biotechnological uses. One of the main yeast antioxidant systems involves two small proteins called thioredoxins. When these two proteins are removed, wine yeast shows diminished growth, protein synthesis, and sugar metabolism under wine-making conditions, and amino acid and lipid metabolism are also affected. Altogether, our results indicate that proper redox regulation is a key factor for metabolic adaptations during grape juice fermentation.

Dataset Information

Saccharomyces cerevisiae ER membrane protein complex subunit 4 (EMC4) plays a crucial role in eIF2B-mediated translation regulation and survival under stress conditions.

Publications

Saccharomyces cerevisiae ER membrane protein complex subunit 4 (EMC4) plays a crucial role in eIF2B-mediated translation regulation and survival under stress conditions.

Similar Datasets

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