IGF-I activates the mouse type IIb myosin heavy chain gene

IGF-I increases skeletal muscle mass, but whether IGF-I increases type IIb myosin heavy chain (MyHC) transcriptional activity is not known. [C.sub.2][C.sub.12] myotubes were cultured with or without IGF-I to determine whether IGF-I increases type IIb MyHC promoter activity, and if so, what region of the promoter might IGF-I signaling regulate. At differentiation days 3 and 4, IGF-I increased type IIb MyHC mRNA and mouse 3.0-kb type IIb MyHC promoter activity. Deletion construct studies identified a potential IGF-I-responsive region between 1.25 and 1.2 kb of the type IIb MyHC promoter, which contained an exact 6-bp T-cell factor/lymphoid enhancer factor (Tcf/Lef) binding site at position--1206 to--1201. Site-specific mutation of the putative Tcf/Lef binding site reduced IGF-I-induced 1.3-kb type IIb MyHC promoter activity. To identify potential 1GF-I signaling molecules, the phosphatidylinositol 3-kinase (PI3K) inhibitors wortmannin and LY-294002 were both found to markedly attenuate IGF-I activation of the 1.3-kb type IIb MyHC promoter. Downstream signaling of IGF-I can phosphorylate and inactivate GSK-3[beta], thereby enhancing [beta]-catenin protein. The GSK-313 inhibitor, LiCI, dramatically enhanced IGF-I induction of the 1.3-kb type IIb MyHC promoter, and constitutively active GSK-3[beta] attenuated IGF-I-induced 1.3-kb type IIb MyHC promoter activity. Finally, IGF-I increased nuclear 13-catenin protein, and small interfering RNA knockdown of [3-catenin attenuated IGFI-induced 1.3-kb type IIb MyHC promoter activity and type IIb MyHC mRNA. In summary, IGF-I stimulation of [C.sub.2][C.sub.12] myotubes increases mouse type IIb MyHC promoter activity, likely through signaling of PI3K, GSK-3[beta], [beta]-catenin, and a Tcf/Lef binding site at -1,206 to -1,201 bp in the promoter. insulin-like growth factor I; [C.sub.2][C.sub.12] myocytes; [beta]-catenin doi: 10.1152/ajpcell.00169.2009