Global studies of eIF5B function in Arabidopsis

The translation initiation factor eIF5B is a GTPase that acts late in the translation initiation process and is one of two universally conserved translation initiation factors. However, the molecular mechanism by which eIF5B is involved in plant development and environmental response remains unknown. In this study, we found that a missense mutation in an eIF5B gene in Arabidopsis thaliana (eIF5B-1; At1g76810) prevents acclimation to high temperature and that severe alleles have multiple growth defects. Surprisingly, Arabidopsis has three other eIF5B genes, none of which is capable of substituting for eIF5B-1 in thermotolerance or growth. One of these other genes, eIF5B-2 (At1g21160) shows differences in level and tissue distribution of expression explaining its inability to complement the eIF5B-1 mutants. The other two genes, eIF5B-3 (At1g76720) and eIF5B-4 (At1g76820) have clearly undergone evolutionary conversion to a non-functional state. The thermotolerance-defective eIF5B-1 allele, a Ser substitution of a highly conserved Gly residue in Domain II, grows normally in the absence of stress. However, polysome profiling and RNA-seq analysis show delayed recovery of translation after heat stress and reduced polysome loading of certain heat shock protein (HSP) mRNAs. A second mutant allele that truncates eIF5B-1 by 30 aa, shows a significantly reduced growth rate. Polysome profiling reveals a dramatic increase in small polysomes and selective changes in polysome loading in the truncation mutant compared to wild type. RNA-seq data suggest that the mutant selectively decreases translation efficiency of auxin and ribosome related genes. We speculate that the phenotypes of these eIF5B-1 mutants result from the requirement for optimal eIF5B activity for translation of specific mRNAs, rather than arising solely from reduced overall translation initiation.