C-terminal Src Kinase (Csk)-mediated Phosphorylation of Eukaryotic Elongation Factor 2 (eEF2) Promotes Proteolytic Cleavage and Nuclear Translocation of eEF2

Yao, Qi; Liu, Bingqian; Li, Hui; McGarrigle, Deirdre; Xing, Bowen; Zhou, Maotian; Wang, Zhe; Zhang, J. Jillian; Huang, Xin‐Yun; Guo, Lin

doi:10.1074/jbc.m113.546481

Cited by 23 publications

(25 citation statements)

References 55 publications

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“…Yao et al proposed C-terminal eEF2 fragments located in the nucleus promote abnormal nuclear morphology and polyploidy [42]. However, we immunohistochemically stained formalin-fixed control and eEF2K -/− cells with an antibody against C-terminal eEF2 and no nuclear staining was observable (Supplementary data).…”

Section: Discussionmentioning

confidence: 89%

Eukaryotic elongation factor 2 is a prognostic marker and its kinase a potential therapeutic target in HCC

et al. 2017

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Hepatocellular carcinoma is a cancer with increasing incidence and largely refractory to current anticancer drugs. Since Sorafenib, a multikinase inhibitor has shown modest efficacy in advanced hepatocellular carcinoma additional treatments are highly needed. Protein phosphorylation via kinases is an important post-translational modification to regulate cell homeostasis including proliferation and apoptosis. Therefore kinases are valuable targets in cancer therapy. To this end we performed 2D differential gel electrophoresis and mass spectrometry analysis of phosphoprotein-enriched lysates of tumor and corresponding non-tumorous liver samples to detect differentially abundant phosphoproteins to screen for novel kinases as potential drug targets. We identified 34 differentially abundant proteins in phosphoprotein enriched lysates. Expression and distribution of the candidate protein eEF2 and its phosphorylated isoform was validated immunohistochemically on 78 hepatocellular carcinoma and non-tumorous tissue samples. Validation showed that total eEF2 and phosphorylated eEF2 at threonine 56 are prognostic markers for overall survival of HCC-patients. The activity of the regulating eEF2 kinase, compared between tumor and non-tumorous tissue lysates by in vitro kinase assays, is more than four times higher in tumor tissues. Functional analyzes regarding eEF2 kinase were performed in JHH5 cells with CRISPR/Cas9 mediated eEF2 kinase knock out. Proliferation and growth is decreased in eEF2 kinase knock out cells.ConclusioneEF2 and phosphorylated eEF2 are prognostic markers for survival of hepatocellular carcinoma patients and the regulating eEF2 kinase is a potential drug target for tumor therapy.

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Section: Discussionmentioning

confidence: 89%

Eukaryotic elongation factor 2 is a prognostic marker and its kinase a potential therapeutic target in HCC

et al. 2017

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“…This notion is largely consistent with previous reports showing that CSK can signal independently from Src. For instance, CSK plays a critical role in mediating G protein signals to actin cytoskeletal reorganization (Lowry et al, 2002); eukaryotic elongation factor 2 (eEF2) phosphorylation induced by CSK promotes proteolytic cleavage and nuclear translocation of eEF2 to induce cell transformation (Yao et al, 2014); and tyrosine phosphorylation of CD45 phosphotyrosine phosphatase by CSK modulates immune signaling (Autero et al, 1994). Besides, regulation of Src-like activity during evolution, such as in the unicellular choanoflagellate Monosiga ovata was not coupled to CSK, suggesting an original Src-independent function for CSK (Segawa et al, 2006).…”

Section: Discussionmentioning

confidence: 99%

Dimerization of the Pragmin pseudo-kinase regulates protein tyrosine phosphorylation

Lecointre

Simon

Kerneur

et al. 2017

Preprint

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The pseudo-kinase and signaling protein Pragmin has been linked to cancer by regulating protein tyrosine phosphorylation via unknown mechanisms. Here we present the crystal structure of the Pragmin 906-1368 amino acids C-terminus, which encompasses its kinase domain. We show that Pragmin contains a classical protein kinase fold devoid of catalytic activity. A particular inhibitory triad, conserved in a Pragmin/SgK269/PEAK1/C19orf35 superfamily, tightly holds the catalytic lysine (K997) to prevent ATP binding. By proteomics, we discovered that this pseudo-kinase uses the tyrosine kinase CSK to induce protein tyrosine phosphorylation in human cells. Interestingly, the protein kinase domain is bordered by N-and C-terminal extensions forming an original dimerization domain that regulates Pragmin self-association and stimulates CSK activity. A1329E mutation in the C-terminal extension destabilizes Pragmin dimerization and reduces CSK activation. Thus, our results reveal a new dimerization mechanism by which a pseudo-kinase can induce protein tyrosine phosphorylation.All rights reserved. No reuse allowed without permission.

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“…Due to its inhibitory role on the pro‐oncogenic SFK members, Csk has been considered a tumor suppressor . However, recent studies suggest that Csk could also contribute to oncogenesis by phosphorylating non‐SFK substrates . Following tyrosine phosphorylation by Csk, eukaryotic elongation factor 2 undergoes SUMOylation and subsequent cleavage, thereby causing chromosomal instability that stimulates neoplastic transformation of cells .…”

mentioning

confidence: 99%

“…However, recent studies suggest that Csk could also contribute to oncogenesis by phosphorylating non‐SFK substrates . Following tyrosine phosphorylation by Csk, eukaryotic elongation factor 2 undergoes SUMOylation and subsequent cleavage, thereby causing chromosomal instability that stimulates neoplastic transformation of cells . Csk also mediates signals generated by the G protein‐coupled receptor, which induces actin stress fiber formation that regulates cell motility independently of SFKs .…”

mentioning

confidence: 99%

“…(18)(19)(20)(21)(22)(23) Following tyrosine phosphorylation by Csk, eukaryotic elongation factor 2 undergoes SUMOylation and subsequent cleavage, thereby causing chromosomal instability that stimulates neoplastic transformation of cells. (22) Csk also mediates signals generated by the G protein-coupled receptor, which induces actin stress fiber formation that regulates cell motility independently of SFKs. (23) More recently, Src was found to act as a negative regulator of Ras, suggesting a stimulatory role of Csk in Ras signaling.…”

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confidence: 99%

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C‐terminal Src kinase‐mediated EPIYA phosphorylation of Pragmin creates a feed‐forward C‐terminal Src kinase activation loop that promotes cell motility

2016

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Pragmin is one of the few mammalian proteins containing the Glu‐Pro‐Ile‐Tyr‐Ala (EPIYA) tyrosine‐phosphorylation motif that was originally discovered in the Helicobacter pylori CagA oncoprotein. Following delivery into gastric epithelial cells by type IV secretion and subsequent tyrosine phosphorylation at the EPIYA motifs, CagA serves as an oncogenic scaffold/adaptor that promiscuously interacts with SH2 domain‐containing mammalian proteins such as the Src homology 2 (SH2) domain‐containing protein tyrosine phosphatase‐2 (SHP2) and the C‐terminal Src kinase (Csk), a negative regulator of Src family kinases. Like CagA, Pragmin also forms a physical complex with Csk. In the present study, we found that Pragmin directly binds to Csk by the tyrosine‐phosphorylated EPIYA motif. The complex formation potentiates kinase activity of Csk, which in turn phosphorylates Pragmin on tyrosine‐238 (Y238), Y343, and Y391. As Y391 of Pragmin comprises the EPIYA motif, Pragmin–Csk interaction creates a feed‐forward regulatory loop of Csk activation. Together with the finding that Pragmin and Csk are colocalized to focal adhesions, these observations indicate that the Pragmin–Csk interaction, triggered by Pragmin EPIYA phosphorylation, robustly stimulates the kinase activity of Csk at focal adhesions, which direct cell‐matrix adhesion that regulates cell morphology and cell motility. As a consequence, expression of Pragmin and/or Csk in epithelial cells induces an elongated cell shape with elevated cell scattering in a manner that is mutually dependent on Pragmin and Csk. Deregulation of the Pragmin–Csk axis may therefore induce aberrant cell migration that contributes to tumor invasion and metastasis.

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C-terminal Src Kinase (Csk)-mediated Phosphorylation of Eukaryotic Elongation Factor 2 (eEF2) Promotes Proteolytic Cleavage and Nuclear Translocation of eEF2

Cited by 23 publications

References 55 publications

Eukaryotic elongation factor 2 is a prognostic marker and its kinase a potential therapeutic target in HCC

Eukaryotic elongation factor 2 is a prognostic marker and its kinase a potential therapeutic target in HCC

Dimerization of the Pragmin pseudo-kinase regulates protein tyrosine phosphorylation

C‐terminal Src kinase‐mediated EPIYA phosphorylation of Pragmin creates a feed‐forward C‐terminal Src kinase activation loop that promotes cell motility

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