Identification of a cellular protein substrate phosphorylated by the avian sarcoma virus-transforming gene product

doi:10.1016/0092-8674(80)90446-8

Cell

Volume 21, Issue 3, October 1980, Pages 829-836

https://doi.org/10.1016/0092-8674(80)90446-8 Get rights and content

Abstract

The avian sarcoma virus-transforming gene product (pp60^src) appears potentially able to mediate cell transformation via phosphorylation since it is tightly associated with a protein kinase activity. We have searched for and have been able to identify a normal cellular protein that appears to be a substrate of pp60^src. The phosphorylation of this protein (34K) is transformation-specific in ASV-transformed cells of both avian and mammalian origin. Moreover, the 34K polypeptide serves as a substrate for the pp60^src phosphotransferase activity in vitro and is phosphorylated at a site identical to the major site of phosphorylation in vivo. These data suggest that upon transformation the 34,000-dalton protein is phosphorylated directly as a result of pp60^src activity.

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The hypervariable N-terminal domain of AnxA2 contains the Tyr-23 and Ser-25 phosphorylation sites as well as the binding site for p11/S100A10, the AnxA2 ligand. Studies have showed that tyrosine 23 can be phosphorylated by pp60src, or after activation of certain other membrane-associated kinases, such as the receptor for insulin, insulin-like growth factor 1 (IGF-1), and platelet derived growth factor (PDGF) (Erikson and Erikson, 1980) (Zhao et al., 2003) (Biener et al., 1996). The phosphorylation of Tyr-23 by Src kinase was proposed to act either as a targeting signal for the plasmamembrane together with p11 or as a negative signal for (AnxA2)2-(p11)2 heterotetramer formation or stabilization (Deora et al., 2004) (He et al., 2008) (Hubaishy et al., 1995) (Zheng et al., 2011).
Colorectal cancer (CRC) is a highly common malignancy, being the third leading cause of cancer death worldwide. Recent epidemiological studies have indicated that carcinogenic effect of diet was mainly attributed to high-fat diets. To investigate the mechanism of high-fat diet-induced colorectal cancer, we systematically quantified the phosphoproteome in human HT-29 cells treated with sodium palmitate (PA). p-Annexin A2 (S26) was predicted to be specifically up-regulated by PA. We confirmed that PA-induced Annexin A2 phosphorylation at Ser26 in C57BL/6 J-Apc^Min/J mice fed with high-fat diet. Phosphorylation of Annexin A2 at Ser26 promotes PA-induced proliferation of HT-29 cells. Moreover, PA suppressed SERCA activity and SERCA2 expression was compensatorily increased. Mechanistically, SERCA2 can partially reverse Annexin A2 phosphorylation at Ser26 caused by PA through intracellular calcium release. Finally, SERCA2 knockdown inhibited high-fat diet-induced tumor growth and Annexin A2 phosphorylation at Ser26 in SCID mice. In all, our studies demonstrate that high-fat diet promotes colorectal carcinogenesis through SERCA2 mediated serine phosphorylation of Annexin A2.
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In addition, two other proteins, Annexin A1 and Lamin, not involved in glycolysis, has been identified by our proteomic analysis. Annexin A1 have been implicated in many cellular processes such as inflammation [26–28], glucorticoid action [29], secretion and exocytosis [30–32], cell growth and differentiation [33–35] and is phosphorylated by several protein tyrosine kinase (PTKs) [36, 37]. As far as Lamin is concerned, its phosphorylation is mainly dependent on the cell cycle [38–40].
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Citation Excerpt :
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The current knowledge of the role of phosphorylation in the functional regulation of AnxA2 is reviewed. To provide a more comprehensive treatment of this topic, we also address in depth the phosphorylation process in general and discuss its possible conformational effects. Furthermore, we discuss the apparent limitations of the methods used to investigate phosphoproteins, as exemplified by the study of AnxA2.
AnxA2 is subjected to complex regulation by post-translational modifications affecting its cellular functions, with Ser11, Ser25 and Tyr23 representing important phosphorylation sites. Thus, Ser phosphorylation of AnxA2 is involved in the recruitment and docking of secretory granules, the regulation of its association with S100A10, and sequestration of perinuclear, translationally inactive mRNP complexes. By contrast, Tyr phosphorylation of AnxA2 regulates its role in actin dynamics and increases its association with endosomal compartments. Modification of its three main phosphorylation sites is not sufficient to discriminate between its numerous functions. Thus, fine-tuning of AnxA2 function is mediated by the joint action of several post-translational modifications.
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Cell

ArticleIdentification of a cellular protein substrate phosphorylated by the avian sarcoma virus-transforming gene product

Abstract

J. Biol. Chem.

Virology

J. Biol. Chem.

Cell

Cell

Surface ruffles as markers for studies of cell transformation by Rous sarcoma virus

Identification of a transformation-specific antigen induced by an avian sarcoma virus

Nature

Fluorographic detection of radioactivity in polyacrylamide gels with the water-soluble fluor, sodium salicylate

Anal. Biochem.

Protein kinase activity associated with the avian sarcoma virus src gene product

Structural analysis of the avian sarcoma virus transforming protein: sites of phosphorylation

J. Virol.

Article
Identification of a cellular protein substrate phosphorylated by the avian sarcoma virus-transforming gene product