Abstract
Insulin activates hexose transport via at least two mechanisms: a p21ras-dependent pathway, leading to an increase in the amount of cell surface GLUT1; and a metabolic, p21ras-independent pathway, leading to translocation of the insulin-responsive transporter GLUT4 to the cell surface. Following insulin stimulation, SHPTP2, a non-transmembrane protein-tyrosine phosphatase, associates with insulin receptor substrate 1 via its Src homology 2 (SH2) domains. Microinjection of a glutathione S-transferase fusion protein encoding the N- and C-terminal SH2 domains of SHPTP2 (GST-NC-SH2) or anti-SHPTP2 antibodies into NIH-3T3 fibroblasts overexpressing the insulin receptor blocks insulin-induced DNA synthesis. Microinjection of either GST-NC-SH2 or anti-SHPTP2 antibodies into 3T3-L1 adipocytes inhibited the insulin-stimulated increase in expression of GLUT1. In contrast, translocation of GLUT4 to the cell surface was unaffected by either GST-NC-SH2 or anti-SHPTP2 antibodies. These data confirm a role for SHPTP2 in insulin-stimulated mitogenesis and indicate that whereas SHPTP2 is necessary for insulin-stimulated expression of GLUT1, it is not required for activation of the metabolic pathway leading to GLUT4 translocation.
Highlights
From the Department of Cell Biology, Harvard Medical School, and the +Molecular Medicine Unit, Beth Israel Hospital, Boston, Massachusetts 02215
Translocation of GLUT4 to the cell surface was unaffected by either glutathione S-transferase (GST)-NC-Src homology 2 (SH2) or anti-SHPTP2 antibodies. These data confirm a role for SHPTP2 in insulin-stimulated mitogenesis and indicate that whereas SHPTP2 is necessary for insulin-stimulated expression of GLUTl, it is not required for activation of the metabolic pathway leading to GLUT4 translocation
We in ves ti gated t he role ofSHPTP2 in th e pathways through wh ich in sulin stimulates hexose uptak e in cultured adipose cells. For these experime nts, two reagents were gen erated for microinj ection into 3T3-LI adipocytes: a glutathione S-transferase (GST) fusion protein enc odin g th e N- and C-te r minal SH2 domains of SHPTP2 (GST-NC-SH2), a nd a ntibodies agai nst full -len gth SHPTP2
Summary
Association of SHPTP2 through its N-terminal SH2 domain with a phosphotyrosyl (pY) peptide corresponding to its binding site on either the PDGF receptor (13) or IRS-l (14, 15) leads to a substantial increase in phosphatase activity. This might well provide a signaling mechanism utilized by SHPTP2 that does not rely on recruitment of a Grb2/S0S complex. Since SHPTP2 is required in both mitogenic and non-mitogenic signaling pathways, we asked whether SHPTP2 is required for the insulin-stimulated increase in cell surface expression of glucose transporters
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