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Translational fidelity and growth of Arabidopsis require stress-sensitive diphthamide biosynthesis

ABSTRACT: Diphthamide, a post-translationally modified histidine residue of eukaryotic TRANSLATION ELONGATION FACTOR2 (eEF2), is the human host cell-sensitizing target of diphtheria toxin. Diphthamide biosynthesis depends on the 4Fe-4S-cluster protein Dph1 catalyzing the first committed step, as well as Dph2 to Dph7, in yeast and mammals. Here we show that diphthamide modification of eEF2 is conserved in Arabidopsis thaliana and requires AtDPH1. Ribosomal −1 frameshifting-error rates are increased in Arabidopsis dph1 mutants, similar to yeast and mice. Compared to the wild type, shorter roots and smaller rosettes of dph1 mutants result from fewer formed cells. TARGET OF RAPAMYCIN (TOR) kinase activity is attenuated, and autophagy is activated, in dph1 mutants. Under abiotic stress diphthamide-unmodified eEF2 accumulates in wild-type seedlings, most strongly upon heavy metal excess, which is conserved in human cells. In summary, our results suggest that diphthamide contributes to the functionality of the translational machinery monitored by plants to regulate growth. Diphthamide is a post-translationally modified histidine residue present in animal and yeast TRANSLATION ELONGATION FACTOR2. Here the authors show that diphthamide modification of eEF2 is conserved in Arabidopsis thaliana and contributes to translational fidelity and growth via cell proliferation.

SUBMITTER: Zhang H

PROVIDER: S-EPMC9273596 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Role of eEF2 diphthamide modification in translational fidelity

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Project description:Role of eEF2 diphthamide modification in translational fidelity

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Project description:Diphthamide biosynthesis involves a carbon-carbon bond-forming reaction catalyzed by a radical S-adenosylmethionine (SAM) enzyme that cleaves a carbon-sulfur (C-S) bond in SAM to generate a 3-amino-3-carboxypropyl (ACP) radical. Using rapid freezing, we have captured an organometallic intermediate with an iron-carbon (Fe-C) bond between ACP and the enzyme's [4Fe-4S] cluster. In the presence of the substrate protein, elongation factor 2, this intermediate converts to an organic radical, formed by addition of the ACP radical to a histidine side chain. Crystal structures of archaeal diphthamide biosynthetic radical SAM enzymes reveal that the carbon of the SAM C-S bond being cleaved is positioned near the unique cluster Fe, able to react with the cluster. Our results explain how selective C-S bond cleavage is achieved in this radical SAM enzyme.

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