Abstract 1225: Gab1-SHP2-ERK Signaling Pathway Comprises an Inhibitory Axis for Insulin-like Growth Factor-1-induced Myogenic Differentiation
Background and Aim: Muscle development requires myoblast differentiation. Most growth factors either stimulate proliferation or induce differentiation of myoblasts. Gab1, a scaffolding/ docking protein, transmits signals from most growth factor receptors mainly to extracellular-signal regulated kinase (ERK) via tyrosine phosphatase, SHP2, and Akt via phosphatidylinositol 3-kinase (PI3-kinase), respectively. We previously reported that Gab1 inhibits 2% horse serum-induced myogenic differentiation of C2C12 myoblast in an SHP2-dependent manner. Insulin-like growh factor (IGF) secreted from myoblasts under 2% horse serum condition is suggested to be critical for myoblast differentiation. Thus, we aimed to investigate how Gab1 regulates IGF-1-induced differentiation of myoblast.
Methods and Results: To delineate Gab1-regulated signaling during myogenesis, we constructed four kinds of adenovirus vectors those carrying wild-type Gab1 (AdGab1WT), mutated Gab1 lacking SHP2 binding site (Gab1DSHP2), mutated Gab1 lacking p85 binding site (AdGab1Dp85) and beta-galactosidase (Adb-gal). IGF-1 induced tyrosine-phosphorylation of Gab1 and complex formation of SHP2 with Gab1. On the other hand, Gab1 associated with p85 PI3-kinase subunit constitutively. Compared with Adb-gal-infected myoblasts, IGF-1-induced myogenic differentiation was enhanced in AdGab1DSHP2-infected myoblasts but inhibited in either AdGab1WT-infected or AdGab1Dp85-infected cells. Conversely, IGF-1-induced ERK activation was inhibited in AdGab1DSHP2-infected cells but enhanced in AdGab1WT or AdGab1Dp85-infected cells. These data suggested that Gab1-SHP2 complex inhibits myogenesis through activating ERK. To test the inhibitory role of ERK for myogenic differentiarion, myoblasts infected with AdGab1WT were pre-treated with or without a MEK inhibitor, U0126, before exposing myoblasts to IGF-1. U0126 reversed inhibitory effect of AdGab1WT and promoted myogenesis. Furthermore, coinfection of AdGab1DSHP2 with a constitutive-active MEK1-expressing adenovirus repressed IGF-1-induced myogeninc differentiation.
Conclusion: Our findings indicate that Gab1-SHP2-ERK signaling pathway comprises an inhibitory axis for IGF-1-induced myogenesis.