Regulation of Insulin Receptor Signaling by the Protein Tyrosine Phosphatase TCPTP

Abstract

The human protein tyrosine phosphatase TCPTP exists as two forms: an endoplasmic reticulum-targeted 48-kDa form (TC48) and a nuclear 45-kDa form (TC45). Although targeted to the nucleus, TC45 can exit in response to specific stimuli to dephosphorylate cytoplasmic substrates. In this study, we investigated the downregulation of insulin receptor (IR) signaling by TCPTP. In response to insulin stimulation, the TC48-D182A and TC45-D182A "substrate-trapping" mutants formed stable complexes with the endogenous tyrosinephosphorylated IR ␤-subunit in 293 cells. Moreover, in response to insulin stimulation, the TC45-D182A mutant accumulated in the cytoplasm of cells overexpressing the IR and in part colocalized with the IR ␤-subunit at the cell periphery. These results indicate that the IR may serve as a cellular substrate for both TC48 and TC45. In immortalized TCPTP ؊/؊ murine embryo fibroblasts, insulin-induced IR ␤-subunit tyrosine phosphorylation and protein kinase PKB/Akt activation were enhanced relative to the values in TCPTP ؉/؉ cells. Importantly, the expression of TC45 or TC48 to physiological levels suppressed the enhanced insulininduced signaling in TCPTP ؊/؊ cells. These results indicate that the differentially localized variants of TCPTP may dephosphorylate the IR and downregulate insulin-induced signaling in vivo.Insulin binding to its cell surface transmembrane receptor stimulates intrinsic protein tyrosine kinase activity, autophosphorylation, and subsequent tyrosyl phosphorylation of insulin receptor substrate (IRS) proteins and adapter proteins such as Shc and Cbl/CAP (reviewed in references 5, 40, 54, and 55). Tyrosyl phosphorylation of IRS and adapter proteins generates docking sites for src homology 2 (SH2) domain-containing signaling proteins such as the p85 subunit of phosphatidylinositol 3-kinase. Collectively, these signaling proteins mediate the biological effects of insulin, which include antilipolysis, glucose uptake, glycogen synthesis, and cell growth (reviewed in references 5, 40, 54, and 55).Protein tyrosine phosphatases (PTPs) have a prominent role in the control of insulin receptor (IR) signaling. PTPs dephosphorylate the IR and its substrates and thus serve to inactivate the IR and terminate signaling. The IR and IRS proteins are rapidly and transiently phosphorylated in response to insulin, with phosphotyrosine (pTyr) content in the IR returning to basal levels within minutes of stimulation (10, 16). It is probable that numerous PTPs participate in the dephosphorylation of the IR and the multiple downstream tyrosine-phosphorylated signaling proteins. Several PTPs have been implicated in the negative regulation of the IR, including the endoplasmic reticulum-associated PTP1B (6,8,49,50) and the transmembrane PTPs LAR (34) and PTP␣ (30,41). PTP␣ has been shown to inhibit insulin-induced prolactin gene expression without altering the phosphorylation status of the IR, IRS-1, or Shc (26). Similarly, LAR may not dephosphorylate the IR in vivo but may downregulate insulin signaling by dephos...

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“…This suggests that, at least in Rat-1 and HepG2 cells, both PTP1B and TC45 function in down-regulation of the signaling response to insulin. Furthermore, in a parallel study, it was shown that in immortalized fibroblasts derived from TC-PTP Ϫ/Ϫ mice the signaling response to insulin was enhanced compared with control TC-PTP ϩ/ϩ fibro- blasts, consistent with our data and the conclusion that, indeed, insulin receptor signaling is regulated by TC-PTP (55).…”

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