Differential regulation of components of the focal adhesion complex by heregulin: Role of phosphatase SHP-2 (original) (raw)
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Scientific reports, 2015
Directed cell migration requires dynamical control of the protein complex within focal adhesions (FAs) and this control is regulated by signaling events involving tyrosine phosphorylation. We screened the SH2 domains present in tyrosine-specific kinases and phosphatases found within FAs, including SRC, SHP1 and SHP2, and examined whether these enzymes transiently target FAs via their SH2 domains. We found that the SRC_SH2 domain and the SHP2_N-SH2 domain are associated with FAs, but only the SRC_SH2 domain is able to be regulated by focal adhesion kinase (FAK). The FAK-dependent association of the SRC_SH2 domain is necessary and sufficient for SRC FA targeting. When the targeting of SRC into FAs is inhibited, there is significant suppression of SRC-mediated phosphorylation of paxillin and FAK; this results in an inhibition of FA formation and maturation and a reduction in cell migration. This study reveals an association between FAs and the SRC_SH2 domain as well as between FAs and ...
Identification of SH2B1 as a focal adhesion protein that regulates focal adhesion size and number
Journal of Cell Science, 2011
The adaptor protein SH2B1b participates in regulation of the actin cytoskeleton during processes such as cell migration and differentiation. Here, we identify SH2B1b as a new focal adhesion protein. We provide evidence that SH2B1b is phosphorylated in response to phorbol 12-myristate 13-acetate (PMA)-induced protein kinase C (PKC) activation and show that PMA induces a rapid redistribution of SH2B1b out of focal adhesions. We also show that growth hormone (GH) increases cycling of SH2B1b into and out of focal adhesions. Ser161 and Ser165 in SH2B1b fall within consensus PKC substrate motifs. Mutating these two serine residues into alanine residues abrogates PMA-induced redistribution of SH2B1b out of focal adhesions, decreases SH2B1b cycling into and out of focal adhesions in control and GH-stimulated cells, and increases the size of focal adhesions. By contrast, mutating Ser165 into a glutamate residue decreases the amount of SH2B1b in focal adhesions and increases the number of focal adhesions per cell. These results suggest that activation of PKC regulates SH2B1b focal adhesion localization through phosphorylation of Ser161 and/or Ser165. The finding that phosphorylation of SH2B1b increases the number of focal adhesions suggests a mechanism for the stimulatory effect on cell motility of SH2B1b.
Cell Growth Differentiation the Molecular Biology Journal of the American Association For Cancer Research, 2000
and is an early event following cell adhesion to fibronectin. In both cases, this regulation is largely dependent on Tyr-397. We also show that the FAK activation loop tyrosines are required for maximal Tyr-397 phosphorylation. Finally, immunostaining analyses revealed that tyrosine-phosphorylated forms of FAK are present in both newly forming and mature focal adhesions. Our findings support a model for reciprocal activation of FAK and Src-family kinases and suggest that FAK/Src signaling may occur during both focal adhesion assembly and turnover. . The abbreviations used are: ECM, extracellular matrix; FAK, focal adhesion kinase; SH2, Src homology 2; WT, wild type.
Molecular and cellular biology, 1999
The coordinated interplay of substrate adhesion and deadhesion is necessary for cell motility. Using MCF-7 cells, we found that insulin-like growth factor I (IGF-I) induces the adhesion of MCF-7 to vitronectin and collagen in a dose- and time-dependent manner, suggesting that IGF-I triggers the activation of different integrins. On the other hand, IGF-I promotes the association of insulin receptor substrate 1 with the focal adhesion kinase (FAK), paxillin, and the tyrosine phosphatase SHP-2, resulting in FAK and paxillin dephosphorylation. Abrogation of SHP-2 catalytic activity with a dominant-negative mutant (SHP2-C>S) abolishes IGF-I-induced FAK dephosphorylation, and cells expressing SHP2-C>S show reduced IGF-I-stimulated chemotaxis compared with either mock- or SHP-2 wild-type-transfected cells. This impairment of cell migration is recovered by reintroduction of a catalytically active SHP-2. Interestingly, SHP-2-C>S cells show a larger number of focal adhesion contacts ...
Journal of Biological Chemistry, 2008
PTPD1 is a cytosolic nonreceptor tyrosine phosphatase and a positive regulator of the Src-epidermal growth factor transduction pathway. We show that PTPD1 localizes along actin filaments and at adhesion plaques. PTPD1 forms a stable complex via distinct molecular modules with actin, Src tyrosine kinase, and focal adhesion kinase (FAK), a scaffold protein kinase enriched at adhesion plaques. Overexpression of PTPD1 promoted cell scattering and migration, short hairpin RNA-mediated silencing of endogenous PTPD1, or expression of PTPD1 mutants lacking either catalytic activity (PTPD1 C1108S ) or the FERM domain (PTPD1 ⌬1-325 ) significantly reduced cell motility. PTPD1 and Src catalytic activities were both required for epidermal growth factor-induced FAK autophosphorylation at its active site and for downstream propagation of ERK1/2 signaling. Our findings demonstrate that PTPD1 is a component of a multivalent scaffold complex nucleated by FAK at specific intracellular sites. By modulating Src-FAK signaling at adhesion sites, PTPD1 promotes the cytoskeleton events that induce cell adhesion and migration.
Protein Phosphatase 1 δ is Associated with Focal Adhesions
Cell Adhesion and Communication, 1998
In all mammalian cells protein phosphatase-1 (PPl) exists in three isofonns, defined as a, y l and 6. Immunofluorescence studies with isofonn-specific antibodies indicated that 6, but not ci or y 1, is enriched at focal adhesions in HeLa cells, fibroblasts, endothelial cells and keratinocytes. This was confirmed also by interference reflection microscopy, which indicated that PP16 was in areas of tight adhesion of the membrane to the extracellular matrix at sites where the microfilament cytoskeleton is organized. In all the cell types so far considered the PP16 in focal adhesions represented only a small aliquot of the total PPlS, which was predominantly localized to the nucleus. The association of PP16 to focal adhesions was confirmed by the coimmunoprecipitation of PP16 with the focal adhesion kinase ~~1 2 5~~~ and with the civ integrin. Comparison between the amount of PPlS associated with focal adhesion proteins and that of PP16 recovered in an anti-PP16 immunoprecipitate confirmed that only a minor amount of the enzyme was associated with the focal adhesions. Since some focal adhesion proteins are phosphorylated on S e r m r , it is likely that PPlS may be involved in the regulation of focal adhesion functions and particularly in the signaling pathway generated by cell-substratum adhesion.