A phosphotyrosine displacement mechanism for activation of Src by PTPα

Department of Molecular Biology, Cornell University, Ithaca, NY 14853, USA.
The EMBO Journal (Impact Factor: 10.43). 03/2000; 19(5):964-78.
Source: PubMed


Protein tyrosine phosphatase alpha (PTPalpha) is believed to dephosphorylate physiologically the Src proto-oncogene at phosphotyrosine (pTyr)527, a critical negative-regulatory residue. It thereby activates Src, and PTPalpha overexpression neoplastically transforms NIH 3T3 cells. pTyr789 in PTPalpha is constitutively phosphorylated and binds Grb2, an interaction that may inhibit PTPalpha activity. We show here that this phosphorylation also specifically enables PTPalpha to dephosphorylate pTyr527. Tyr789-->Phe mutation abrogates PTPalpha-Src binding, dephosphorylation of pTyr527 (although not of other substrates), and neoplastic transformation by overexpressed PTPalpha in vivo. We suggest that pTyr789 enables pTyr527 dephosphorylation by a pilot binding with the Src SH2 domain that displaces the intramolecular pTyr527-SH2 binding. Consistent with model predictions, we find that excess SH2 domains can disrupt PTPalpha-Src binding and can block PTPalpha-mediated dephosphorylation and activation in proportion to their affinity for pTyr789. Moreover, we show that, as predicted by the model, catalytically defective PTPalpha has reduced Src binding in vivo. The displacement mechanism provides another potential control point for physiological regulation of Src-family signal transduction pathways.

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    • "However, more remains to be learned about how E-cadherin might activate the RPTPα-SFK pathway. RPTPα can be activated by several mechanisms (den Hertog et al., 1995; Tracy et al., 1995; Zheng et al., 2000), but an interesting clue lies in the capacity for homodimerization to inhibit the catalytic activity of RPTPα. It has been shown recently that disruption of this autoinhibitory mechanism may increase Src activity in a significant subset of colon, breast, and liver human tumors (Huang et al., 2011). "

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    • "RPTPα can be activated by several mechanisms (den Hertog et al., 1995; Tracy et al., 1995; Zheng et al., 2000), but an interesting clue lies in the capacity for homodimerization to inhibit the catalytic activity of RPTPα. It has been shown recently that disruption of this auto-inhibitory mechanism may increase Src activity in significant subset of colon, breast and liver human tumors (Huang et al., 2011). "
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    ABSTRACT: Cell-cell adhesion couples the contractile cortices of epithelial cells together, generating tension to support a range of morphogenetic processes. E-cadherin adhesion plays an active role in generating junctional tension, by promoting actin assembly and cortical signaling pathways that regulate Myosin II. Multiple Myosin II paralogs accumulate at mammalian epithelial cell-cell junctions. Earlier we found that Myosin IIA responds to Rho-ROCK signaling to support junctional tension in MCF-7 cells. Although Myosin IIB is also found at the zonula adherens (ZA) in these cells, its role in junctional contractility, and its mode of regulation, are less well understood. We now demonstrate that Myosin IIB contributes to tension at the epithelial ZA. Further, we identify a RPTPα-SFK-Rap1 pathway as responsible for recruiting Myosin IIB to the ZA and supporting contractile tension. Overall, these findings reinforce the concept that orthogonal E-cadherin-based signaling pathways recruit distinct Myosin II paralogs to generate the contractile apparatus at apical epithelial junctions. © 2015 by The American Society for Cell Biology.
    Molecular Biology of the Cell 01/2015; 26(7). DOI:10.1091/mbc.E14-07-1223 · 4.47 Impact Factor
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    • "Cells were cultured in DMEM supplemented with 10% fetal bovine serum. Genetically modified NIH3T3 fibroblasts that express HA-tagged wild-type PTPα (NIH3T3PTPα) and Y789F unphosphorylatable mutant PTPα (NIH3T3Y789F) under control of a doxycycline sensitive repressor were obtained from Dr. David Shalloway (Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY) and were generated as previously described [20]. These cells were grown in Dulbecco’s modified Eagle medium containing 5% fetal bovine serum in the presence of 5 ng/ml doxycycline. "
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    ABSTRACT: IL-1β contributes to connective tissue destruction in part by up-regulating stromelysin-1 (MMP-3), which in fibroblasts is a focal adhesion-dependent process. Protein tyrosine phosphatase-α (PTPα) is enriched in and regulates the formation of focal adhesions, but the role of PTPα in connective tissue destruction is not defined. We first examined destruction of periodontal connective tissues in adult PTPα(+/+) and PTPα(-/-) mice subjected to ligature-induced periodontitis, which increases the levels of multiple cytokines, including IL-1β. Three weeks after ligation, maxillae were processed for morphometry, micro-computed tomography and histomorphometry. Compared with unligated controls, there was ∼1.5-3 times greater bone loss as well as 3-fold reduction of the thickness of the gingival lamina propria and 20-fold reduction of the amount of collagen fibers in WT than PTPα(-/-) mice. Immunohistochemical staining of periodontal tissue showed elevated expression of MMP-3 at ligated sites. Second, to examine mechanisms by which PTPα may regulate matrix degradation, human MMP arrays were used to screen conditioned media from human gingival fibroblasts treated with vehicle, IL-1β or TNFα. Although MMP-3 was upregulated by both cytokines, only IL-1β stimulated ERK activation in human gingival fibroblasts plated on fibronectin. TIRF microscopy and immunoblotting analyses of cells depleted of PTPα activity with the use of various mutated constructs or with siRNA or PTPα(KO) and matched wild type fibroblasts were plated on fibronectin to enable focal adhesion formation and stimulated with IL-1β. These data showed that the catalytic and adaptor functions of PTPα were required for IL-1β-induced focal adhesion formation, ERK activation and MMP-3 release. We conclude that inflammation-induced connective tissue degradation involving fibroblasts requires functionally active PTPα and in part is mediated by IL-1β signaling through focal adhesions.
    PLoS ONE 08/2013; 8(8):e70659. DOI:10.1371/journal.pone.0070659 · 3.23 Impact Factor
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