Protein Tyrosine Phosphatase PTP20 Induces Actin Cytoskeleton Reorganization by Dephosphorylating p190 RhoGAP in Rat Ovarian Granulosa Cells Stimulated with Follicle-Stimulating Hormone

Gene Research Center, University of Tsukuba, Ibaraki 305-8572, Japan.
Molecular Endocrinology (Impact Factor: 4.02). 05/2003; 17(4):534-49. DOI: 10.1210/me.2002-0187
Source: PubMed


We identified 25 protein tyrosine phosphatases (PTPs) expressed in rat ovarian granulosa cells. Of these PTPs, the expression levels of at least PTP20, PTP-MEG1, PTPepsilonM, and PTPepsilonC significantly changed during the estrous cycle. We examined the cellular functions of PTP20 in granulosa cells by expressing the wild type, a catalytically inactive CS mutant in which Cys229 of PTP20 was changed to Ser, or a substrate-trapping DA mutant in which Asp197 was mutated to Ala, using an adenovirus vector. Overexpression of the wild type, but not of the CS mutant, induced retraction of the cell body with the extension of long, dendritic-like processes after stimulation with FSH, a critical factor for the survival and differentiation of these cells. In addition, cell adhesion to the substratum decreased in an FSH-dependent manner. Inhibiting Rho GTPase activity with C3 botulinum toxin caused similar morphological changes. The FSH-enhanced phosphotyrosine (p-Tyr) level of p190 RhoGAP was selectively reduced by the overexpressed wild type, but not by mutated PTP20. Although p190 RhoGAP is tyrosine phosphorylated by c-Src via the tyrosine kinase Pyk2, wild-type PTP20 had little effect on p-Tyr418 of c-Src and no effect on p-Tyr402 of Pyk2, which are required for full c-Src activity and for interacting between Pyk2 and c-Src, respectively. The CS and DA mutants as well as the wild type reduced the formation of p190 RhoGAP-p120 RasGAP complexes. Confocal microscopy analysis revealed that PTP20 intracellularly colocalizes with p190 RhoGAP. These results demonstrate that PTP20 regulates the functions of granulosa cells in an FSH-dependent manner by dephosphorylating p190 RhoGAP and subsequently inducing reorganization of the actin cytoskeleton. Moreover, our data suggest that PTPs play significant roles in controlling the dynamics of ovarian functions.

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    • "First, it is involved in the negative feedback mechanisms controlling the Bcr-Abl fusion tyrosine kinase-signalling network by inhibiting the phosphorylation of Src family kinase (Rubbi et al, 2011). Second, PTPN18 can induce actin cytoskeleton reorganisation (Shiota et al, 2003), suggesting a plausible explanation of the morphological changes observed in the imatinib-resistant GIST cells (Mahadevan et al, 2007). Third, altered phosphorylation of tyrosine kinases is an alternative mechanism of imatinib resistance in GIST (Takahashi et al, 2013), suggesting the possible involvements of protein phosphatases in imatinib resistance of GIST. "
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    • "Cells were lysed in RIPA buffer, and proteins were isolated from cell lysates by immunoprecipitation as described previously [28]. For immunoblotting, proteins were separated on SDS-polyacrylamide gels under reducing conditions, followed by electrophoretic transfer to PVDF membranes as described previously [29]. "
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    • "The phosphorylation of PAX generates a binding site for p120RasGAP displacing it from its binding partner p190RhoGAP, that once freed may contribute to inhibit RhoA activity [154]. A similar control mechanism operated on p190RhoGAP has been described also for PYK2/Src in rat ovarian granulosa cells and is contrasted by the activity of PTP20 [155]. However the correlation between RhoA and FAK is more complex. "
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