GTP binding to translation factor eIF2B stimulates its guanine nucleotide exchange activity

Christopher J Kershaw,Martin D Jennings,Francesco Cortopassi,Margherita Guaita,Hawra Al-Ghafli,Graham D Pavitt

doi:10.1016/j.isci.2021.103454

Christopher J Kershaw, Martin D Jennings + Show 4 more

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https://doi.org/10.1016/j.isci.2021.103454

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Journal: iScience	Publication Date: Nov 14, 2021
Citations: 7	License type: cc-by

Affiliation: University of Manchester

Abstract

SummaryeIF2B is the guanine nucleotide exchange factor (GEF) required for cytoplasmic protein synthesis initiation in eukaryotes and its regulation within the integrated stress response (ISR). It activates its partner factor eIF2, thereby promoting translation initiation. Here we provide evidence through biochemical and genetic approaches that eIF2B can bind directly to GTP and this can enhance its rate of GEF activity toward eIF2–GDP in vitro. GTP binds to a subcomplex of the eIF2Bγ and ε subunits. The eIF2Bγ amino-terminal domain shares structural homology with hexose sugar phosphate pyrophosphorylase enzymes that bind specific nucleotides. A K66R mutation in eIF2Bγ is especially sensitive to guanine or GTP in a range of functional assays. Taken together, our data suggest eIF2Bγ may act as a sensor of purine nucleotide availability and thus modulate eIF2B activity and protein synthesis in response to fluctuations in cellular nucleotide levels.

Highlights

Translation initiation is a highly regulated phase of gene expression
Several other initiation factors assist as part of a preinitiation complex (PIC), which binds to mRNA near its 50 cap and scans to an AUG codon where codon-anticodon pairing signals selection of the start site
For continued rounds of initiation, the relatively stable eIF2-GDP complex must be reactivated into eIF2GTP for Met-tRNAi binding. This recycling of eIF2 is performed by the guanine nucleotide exchange factor (GEF) eIF2B

Summary

Introduction

Translation initiation is a highly regulated phase of gene expression. Methionine initiator tRNA (Met-tRNAi) is brought to the small ribosomal subunit (40S) as part of a ternary complex (TC) with the GTP-binding translation initiation factor eIF2 in complex with GTP. For continued rounds of initiation, the relatively stable eIF2-GDP complex must be reactivated into eIF2GTP for Met-tRNAi binding. This recycling of eIF2 is performed by the guanine nucleotide exchange factor (GEF) eIF2B. Additional kinases (i.e. PKR, PERK, and HRI) respond to a wider range of cellular stresses (Pavitt, 2018; Wek, 2018). This phosphorylation at a common serine (ser 52) increases the affinity of eIF2 for eIF2B, preventing nucleotide exchange on eIF2 by eIF2B. The reduction in TC levels brings about large-scale reductions in overall protein synthesis rates and translational activation of specific genes that causes wider reprogramming of gene expression in what is termed the integrated stress response (ISR) (Pavitt, 2018; Wek, 2018)

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