Dph3, a Small Protein Required for Diphthamide Biosynthesis, Is Essential in Mouse Development

Liu, Shihui; Wiggins, Jason F.; Sreenath, Taduru; Kulkarni, Ashok B.; Ward, Jerrold M.; Leppla, Stephen H.

doi:10.1128/mcb.26.10.3835-3841.2006

Cited by 66 publications

(70 citation statements)

References 20 publications

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“…In support of this notion, evidence from research groups including our own has shown that diphthamide defects increase translational frame-shifting (Ortiz et al, 2006;Bär et al, 2008). Moreover, homologues of diphthamide synthesis genes (DPH1/OVCA1 and DPH3/KTI11) affect the proliferation and development of mammalian cells, which is why inactivation of DPH3/KTI11 is associated with tRNA modification defects and neurodegeneration and mutations in DPH1/OVCA1 revealed a tumour suppressor role for this diphthamide synthesis gene in ovarian cancer (Chen & Behringer, 2004;Nobukuni et al, 2005;Huang et al, 2005;Liu et al, 2006;Kim et al, 2010 Whether or not this implies structural or regulatory roles for diphthamide in mRNA translation remains to be seen. The latter, however, is intriguing with the emergence of a cellular ADP-ribosyltransferase that resembles the diphthamide-dependent ADPribosylation reaction by DT (Lee & Iglewski, 1984;Jäger et al, 2011).…”

Section: Biological Significance For Diphthamide Modification Of Ef2mentioning

confidence: 75%

“…In the budding yeast Saccharomyces cerevisiae, diphthamide biosynthesis requires at least five genes, DPH1-DPH5 (Liu et al, 2004), two mammalian homologues of which (DPH1/OVCA1 & DPH3/KTI11) are intriguingly involved in embryonic development and cell proliferation in rodents and humans (Fichtner & Schaffrath, 2002;Chen & Behringer, 2004;Fichtner et al, 2003;Liu & Leppla, 2003;Nobukuni et al, 2005;Liu et al, 2006). Though complex in nature, the diphthamide pathway has been shown to be molecularly dissectable.…”

Section: Posttranslational Biosynthesis Of Diphthamide On Ef2mentioning

confidence: 99%

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Diphtheria Disease and Genes Involved in Formation of Diphthamide, Key Effector of the Diphtheria Toxin

Uthman¹,

Liu²,

Giorgini³

et al. 2012

Insight and Control of Infectious Disease in Global Scenario

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Section: Biological Significance For Diphthamide Modification Of Ef2mentioning

confidence: 75%

Section: Posttranslational Biosynthesis Of Diphthamide On Ef2mentioning

confidence: 99%

Diphtheria Disease and Genes Involved in Formation of Diphthamide, Key Effector of the Diphtheria Toxin

Uthman¹,

Liu²,

Giorgini³

et al. 2012

Insight and Control of Infectious Disease in Global Scenario

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“…OVCA1 −/− and Dph4 −/− mice phenocopied each other with a delay in embryo development starting as early as E8.5, and having 100% prenatal lethality (20)(21)(22) (Table S2). Interestingly, the eEF2 G717R/G717R mice displayed a milder phenotype, with a small fraction surviving to adulthood.…”

Section: Resultsmentioning

confidence: 99%

“…Diphthamide deficiency can be induced by deletion of the genes involved in the diphthamide biosynthesis (20)(21)(22). However, these genes may have pleiotropic functions, as seen with Dph3 involvement in tRNA anticodon modifications (23).…”

Section: Resultsmentioning

confidence: 99%

“…Surprisingly, none of the diphthamide-deficient mutants identified in either yeast or CHO cells exhibit strong phenotypes (5, 6, 8-10, 18, 19), demonstrating that the diphthamide modification is not essential for cell survival. However, recent studies of OVCA1 (Dph1), Dph3, and Dph4 knockout mice have shed light on the physiological role of diphthamide in multicellular organisms (20)(21)(22). Remarkably, OVCA1-, Dph3-, and Dph4-null mice die during embryonic development, suggesting an important role of diphthamide in embryonic development.…”

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confidence: 99%

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Diphthamide modification on eukaryotic elongation factor 2 is needed to assure fidelity of mRNA translation and mouse development

Liu

Bachran

Gupta

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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To study the role of the diphthamide modification on eukaryotic elongation factor 2 (eEF2), we generated an eEF2 Gly 717 Arg mutant mouse, in which the first step of diphthamide biosynthesis is prevented. Interestingly, the Gly 717 -to-Arg mutation partially compensates the eEF2 functional loss resulting from diphthamide deficiency, possibly because the added +1 charge compensates for the loss of the +1 charge on diphthamide. Therefore, in contrast to mouse embryonic fibroblasts (MEFs) from OVCA1 −/− mice, eEF2 G717R/G717R MEFs retain full activity in polypeptide elongation and have normal growth rates. Furthermore, eEF2 G717R/G717R mice showed milder phenotypes than OVCA1 −/− mice (which are 100% embryonic lethal) and a small fraction survived to adulthood without obvious abnormalities. Moreover, eEF2 G717R/G717R /OVCA1 −/− double mutant mice displayed the milder phenotypes of the eEF2 G717R/G717R mice, suggesting that the embryonic lethality of OVCA1 −/− mice is due to diphthamide deficiency. We confirmed that the diphthamide modification is essential for eEF2 to prevent −1 frameshifting during translation and show that the Gly 717 -to-Arg mutation cannot rescue this defect. E ukaryotic elongation factor 2 (eEF2) is a member of the GTPbinding translation elongation factor family, and an essential factor for protein synthesis and cell survival. eEF2 drives the GTP-dependent translocation of the nascent polypeptide chain from the A site to the P site of the ribosome and advances mRNA by three bases during the elongation cycle of protein synthesis (1). eEF2 is highly homologous in all eukaryotes. In fact, eEF2 of humans, rats, mice, hamsters, and other mammals have exactly the same amino acid sequence. Intriguingly, all eukaryotic eEF2 proteins contain a unique posttranslationally modified histidine residue termed diphthamide (2, 3). Diphthamide modification occurs after eEF2 is translated and is irreversible, marking the completion of the biosynthesis of eEF2.Although the physiological role of the diphthamide modification on eEF2 remains elusive, diphthamide is the well-known target for the adenosine diphosphate (ADP)-ribosylating toxins from bacterial pathogens, such as diphtheria toxin (DT) from Corynebacterium diphtheriae, Pseudomonas exotoxin A (ETA) from Pseudomonas aeruginosa, and the recently identified cholix toxin (CT) from Vibrio cholerae (4). As virulence factors, these ADP-ribosylating toxins catalyze transfer of the ADP ribose from nicotinamide adenine dinucleotide (NAD + ) to diphthamide on eEF2 (Fig. S1), thus inactivating eEF2, halting cellular protein synthesis, and causing cell death.Because the diphthamide modification is required for the action of the ADP-ribosylating toxins, the complex diphthamide biosynthesis pathway is amenable to genetic analysis, and mutants defective in diphthamide biosynthesis have been isolated in both Chinese hamster ovary (CHO) cells and yeast (Saccharomyces cerevisiae) by selection for resistance to DT or an engineered ADP-ribosylating toxin − anthrax protective a...

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Proteomics analysis of in vitro protein methylation during Src‐induced transformation

Chiou¹,

Fu²,

Lin³

et al. 2012

Electrophoresis

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Src, a nonreceptor tyrosine kinase, was the first oncogene identified from an oncogenic virus. Mechanistic studies of Src-induced transformations aid in understanding the pathologic processes underlying tumorigenesis and may provide new strategies for cancer therapy. Although several pathways and protein modifications are reportedly involved in Src-induced transformation, the detailed mechanisms of their regulation remain unclear. Protein methylation is an important PTM that is widely involved in cellular physiology. In this study, we determined if protein methylation was involved in Src activation and which methylated proteins were associated with this activity. Using in vitro methylation and 2-DE analysis of viral Src (v-Src)-transformed rat kidney epithelial cells (RK3E), several known and novel methylated proteins were identified based on their changes in methylation signal intensity upon transformation. Among these, elongation factor 2 (EF-2), heterogeneous nuclear ribonucleoprotein K (hnRNP K), and β-tubulin protein expressions remained unchanged, indicating that their altered methylation levels were due to Src activation. In addition, the altered expression of β-actin, vimentin, and protein phosphatase 2, catalytic subunit (PPP2C) as well as protein phosphatase 2, catalytic subunit methylation were also confirmed in RK3E cells transformed with a human oncogenic Src mutant (Src531), supporting their association with Src-induced transformation in human cancer. Together, we showed putative involvement of protein methylation in Src activation and our identification of methylated proteins provides important targets for extensively studying Src-induced transformations.

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Dph3, a Small Protein Required for Diphthamide Biosynthesis, Is Essential in Mouse Development

Cited by 66 publications

References 20 publications

Diphtheria Disease and Genes Involved in Formation of Diphthamide, Key Effector of the Diphtheria Toxin

Diphtheria Disease and Genes Involved in Formation of Diphthamide, Key Effector of the Diphtheria Toxin

Diphthamide modification on eukaryotic elongation factor 2 is needed to assure fidelity of mRNA translation and mouse development

Proteomics analysis of in vitro protein methylation during Src‐induced transformation

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