Abstract
The Src homology 2 domain-containing protein-tyrosine phosphatase Shp2 has been implicated in a variety of growth factor signaling pathways, but its role in insulin signaling has remained unresolved. In vitro studies suggest that Shp2 is both a negative and positive regulator of insulin signaling, although its physiological function in a number of peripheral insulin-responsive tissues remains unknown. To address the metabolic role of Shp2 in the liver, we generated mice with either chronic or acute hepatic Shp2 deletion using tissue-specific Cre-LoxP and adenoviral Cre approaches, respectively. We then analyzed insulin sensitivity, glucose tolerance, and insulin signaling in liver-specific Shp2-deficient and control mice. Mice with chronic Shp2 deletion exhibited improved insulin sensitivity and increased glucose tolerance compared with controls. Acute Shp2 deletion yielded comparable results, indicating that the observed metabolic effects are directly caused by the lack of Shp2 in the liver. These findings correlated with, and were most likely caused by, direct dephosphorylation of insulin receptor substrate (IRS)1/2 in the liver, accompanied by increased PI3K/Akt signaling. In contrast, insulin-induced ERK activation was dramatically attenuated, yet there was no effect on the putative ERK site on IRS1 (Ser(612)) or on S6 kinase 1 activity. These studies show that Shp2 is a negative regulator of hepatic insulin action, and its deletion enhances the activation of PI3K/Akt pathway downstream of the insulin receptor.
Highlights
Upon binding to the insulin receptor (IR),7 insulin induces trans-phosphorylation of several tyrosyl residues, leading to the recruitment and phosphorylation of insulin receptor substrates (IRSs) and Grb2associated binder (Gab) family proteins. These serve as docking sites for Src homology 2 domain-containing signal relay molecules, such as Src homology phosphatase 2 (Shp2) and the p85 regulatory subunit of phosphatidylinositol 3-kinase (PI3K)
Shp2 plays an essential role in most receptor tyrosine kinase signaling pathways, where it is required for normal activation of the ERK pathway and its transcriptional targets [10, 11]
In response to IR activation, Shp2 binds to IRS1/2, Gab1, as well as the transmembrane protein Shps1/Sirp␣ [12]
Summary
Mouse Studies—Shp2-floxed (Shp2fl/fl) mice were generated previously [24]. Albumin-Cre mice were obtained from Dr CR. Fed glucose measurements were taken between 7 and 9 a.m. and, where indicated, from mice fasted for 12 h. Biochemical Analyses—For insulin signaling experiments, 20-week-old male mice were fasted overnight, injected intraperitoneally with insulin (10 milliunits/g of body weight), and killed 5 or 10 min after injection. Immunoblot analysis of liver lysates indicated that Shp protein expression was decreased by ϳ50% in HET and ϳ85% in LSHKO mice compared with control mice (Fig. 1, A and B). These findings are consistent with complete deletion of Shp in hepatocytes; the residual Shp in LSHKO liver lysates likely reflects Shp expression in other cell types in the liver, such as biliary cannalicular. Statistical protein-tyrosine phosphatase (PTP) known to regulate hepatic insulin signaling, protein-tyrosine phosphatase 1B (PTP1B) [28, 29], was unaffected in LSHKO mice (Fig. 1A, bottom analysis was performed using unpaired, two-tailed Student’s t-test. * indicates difference between control and LSHKO mice (*, p Յ 0.05; **, p Յ 0.01)
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