Eukaryotic initiation factor 2B, eIF2B is a guanine nucleotide exchange, factor with a central role in coordinating the initiation of translation. During stress and disease, the activity of eIF2B is inhibited via the phosphorylation of its substrate eIF2 (p-eIF2α). A number of different kinases respond to various stresses leading to the phosphorylation of the alpha subunit of eIF2 and collectively this regulation is known as the Integrated Stress Response, ISR. This targeting of eIF2B allows the cell to regulate protein synthesis and reprogramme gene expression to restore homeostasis. Advances within structural biology have furthered our understanding of how eIF2B interacts with eIF2 in both the productive GEF active form and the non-productive eIF2ɑ phosphorylated form. Here, current knowledge of the role of eIF2B in the ISR is discussed within the context of normal and disease states focussing particularly on diseases such as Vanishing White Matter Disease (VWMD) and Permanent Neonatal Diabetes Mellitus (PNDM), which are directly linked to mutations in eIF2B. The role of eIF2B in synaptic plasticity and memory formation is also discussed. In addition, the cellular localisation of eIF2B is reviewed and considered along with the role of additional in vivo eIF2B binding factors and protein modifications that may play a role in modulating eIF2B activity during health and disease.
Eukaryotic initiation factor 2B (eIF2B) is a master regulator of translation control. eIF2B recycles inactive eIF2-GDP to active eIF2-GTP. Under transient/acute cellular stress, a family of kinases phosphorylate the alpha subunit of eIF2 (eIF2α-P[S51]) activating the integrated stress response (ISR). This response pathway inhibits eIF2B activity resulting in overall translation attenuation and reprogramming of gene expression to overcome the stress. The duration of an ISR programme can dictate cell fate wherein chronic activation has pathological outcomes. Vanishing white matter disease (VWMD) is a chronic ISR-related disorder linked to mutations in eIF2B. eIF2B is vital to all cell types, yet VWMD eIF2B mutations primarily affect astrocytes and oligodendrocytes suggesting cell type-specific functions of eIF2B. Regulation of the cytoplasmic localisation of eIF2B (eIF2B bodies) has been implicated in the ISR. Here, we highlight the cell type specific localisation of eIF2B within neuronal and glial cell types. Our analyses revealed that each cell type possesses its own steady-state repertoire of eIF2B bodies with varied subunit composition and activity. We also demonstrate that neural and glial cell types respond similarly to acute induction of the ISR whilst a chronic ISR programme exerts cell type-specific differences. Regulatory composition of eIF2B bodies is suggested to be differentially modulated in a manner that correlates to the action of acute and chronic ISR. We also highlight a cell type specific response of the ISR inhibitor ISRIB on eIF2B localisation and activity.
The multisubunit eukaryotic initiation factor 2B (eIF2B), a guanine nucleotide exchange factor (GEF) for eIF2, is an essential regulator of translation initiation. Activation of the cellular integrated stress response (ISR) by factors such as endoplasmic reticulum stress leads to phosphorylation of eIF2α and inhibition of eIF2B GEF activity. Cytoplasmic bodies containing eIF2B subunits, termed eIF2B bodies, have been shown to alter in subunit composition and fluorescence recovery after photobleaching activity in response to the ISR. Analysis of the subunit composition of endogenous eIF2B bodies is dependent on accurate detection of each protein in a cellular context via immunocytochemistry (ICC). We describe bioinformatic techniques to optimize the ICC detection of eIF2B foci in U373 cells. The screening of commercially available primary antibodies against predicted epitopes enhanced measurements of the number, size and fluorescence intensity of eIF2B bodies. A consistent and reproducible ICC analysis of endogenous eIF2B bodies will aid characterisation of eIF2B bodies during the ISR or under disease conditions.
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