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Analysis of eIF2B bodies and their relationships with stress granules and P-bodies.

ABSTRACT: Eukaryotic cells respond to stress and changes in the environment in part by repressing translation and forming cytoplasmic assemblies called stress granules and P-bodies, which harbor non-translating mRNAs and proteins. A third, but poorly understood, assembly called the eIF2B body can form and contains the eIF2B complex, an essential guanine exchange factor for the translation initiation factor eIF2. Hypomorphic EIF2B alleles can lead to Vanishing White Matter Disease (VWMD), a leukodystrophy that causes progressive white matter loss. An unexplored question is how eIF2B body formation is controlled and whether VWMD alleles in EIF2B alter the formation of eIF2B bodies, stress granules, or P-bodies. To examine these issues, we assessed eIF2B body, stress granule, and P-body induction in wild-type yeast cells and cells carrying VWMD alleles in the EIF2B2 (GCD7) and EIF2B5 (GCD6) subunits of eIF2B. We demonstrate eIF2B bodies are rapidly and reversibly formed independently of stress granules during acute glucose deprivation. VWMD mutations had diverse effects on stress-induced assemblies with some alleles altering eIF2B bodies, and others leading to increased P-body formation. Moreover, some VWMD-causing mutations in GCD7 caused hyper-sensitivity to chronic GCN2 activation, consistent with VWMD mutations causing hyper-sensitivity to eIF2? phosphorylation and thereby impacting VWMD pathogenesis.

SUBMITTER: Moon SL

PROVIDER: S-EPMC6095920 | biostudies-literature | 2018 Aug

REPOSITORIES: biostudies-literature

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Analysis of eIF2B bodies and their relationships with stress granules and P-bodies.

Moon Stephanie L SL Parker Roy R

Scientific reports 20180816 1

Eukaryotic cells respond to stress and changes in the environment in part by repressing translation and forming cytoplasmic assemblies called stress granules and P-bodies, which harbor non-translating mRNAs and proteins. A third, but poorly understood, assembly called the eIF2B body can form and contains the eIF2B complex, an essential guanine exchange factor for the translation initiation factor eIF2. Hypomorphic EIF2B alleles can lead to Vanishing White Matter Disease (VWMD), a leukodystrophy ...[more]

PMID: 30115954

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Project description:Transcriptome analyses indicate that a core 10%-15% of the yeast genome is modulated by a variety of different stresses. However, not all the induced genes undergo translation, and null mutants of many induced genes do not show elevated sensitivity to the particular stress. Elucidation of the RNA lifecycle reveals accumulation of non-translating mRNAs in cytoplasmic granules, P-bodies, and stress granules for future regulation. P-bodies contain enzymes for mRNA degradation; under stress conditions mRNAs may be transferred to stress granules for storage and return to translation. Protein degradation by the ubiquitin-proteasome system is elevated by stress; and here we analyzed the steady state levels, decay, and subcellular localization of the mRNA of the gene encoding the F-box protein, UFO1, that is induced by stress. Using the MS2L mRNA reporter system UFO1 mRNA was observed in granules that colocalized with P-bodies and stress granules. These P-bodies stored diverse mRNAs. Granules of two mRNAs transported prior to translation, ASH1-MS2L and OXA1-MS2L, docked with P-bodies. HSP12 mRNA that gave rise to highly elevated protein levels was not observed in granules under these stress conditions. ecd3, pat1 double mutants that are defective in P-body formation were sensitive to mRNAs expressed ectopically from strong promoters. These highly expressed mRNAs showed elevated translation compared with wild-type cells, and the viability of the mutants was strongly reduced. ecd3, pat1 mutants also exhibited increased sensitivity to different stresses. Our interpretation is that sequestration of highly expressed mRNAs in P-bodies is essential for viability. Storage of mRNAs for future regulation may contribute to the discrepancy between the steady state levels of many stress-induced mRNAs and their proteins. Sorting of mRNAs for future translation or decay by individual cells could generate potentially different phenotypes in a genetically identical population and enhance its ability to withstand stress.

Project description:Eukaryotic translation initiation factor 4E (eIF4E) plays a pivotal role in the control of cap-dependent translation initiation, modulates the fate of specific mRNAs, occurs in processing bodies (PBs) and is required for formation of stress granules (SGs). In this study, we focused on the subcellular localization of a representative compendium of eIF4E protein isoforms, particularly on the less studied members of the human eIF4E protein family, eIF4E2 and eIF4E3.We showed that unlike eIF4E1, its less studied isoform eIF4E3_A, encoded by human chromosome 3, localized to stress granules but not PBs upon both heat shock and arsenite stress. Furthermore, we found that eIF4E3_A interacts with human translation initiation factors eIF4G1, eIF4G3 and PABP1 in vivo and sediments into the same fractions as canonical eIF4E1 during polysome analysis in sucrose gradients. Contrary to this finding, the truncated human eIF4E3 isoform, eIF4E3_B, showed no localization to SGs and no binding to eIF4G. We also highlighted that eIF4E2 may exhibit distinct functions under different stresses as it readily localizes to P-bodies during arsenite and heat stresses, whereas it is redirected to stress granules only upon heat shock. We extended our study to a number of protein variants, arising from alternative mRNA splicing, of each of the three eIF4E isoforms. Our results surprisingly uncovered differences in the ability of eIF4E1_1 and eIF4E1_3 to form stress granules in response to cellular stresses.Our comparison of all three human eIF4E isoforms and their protein variants enriches the intriguing spectrum of roles attributed to the eukaryotic initiation translation factors of the 4E family, which exhibit a distinctive localization within different RNA granules under different stresses. The localization of eIF4E3_A to stress granules, but not to processing bodies, along with its binding to eIF4G and PABP1 suggests a role of human eIF4E3_A in translation initiation rather than its involvement in a translational repression and mRNA decay and turnover. The localization of eIF4E2 to stress granules under heat shock but not arsenite stress indicates its distinct function in cellular response to these stresses and points to the variable protein content of SGs as a consequence of different stress insults.

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Analysis of eIF2B bodies and their relationships with stress granules and P-bodies.

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Analysis of eIF2B bodies and their relationships with stress granules and P-bodies.

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