TableEdit

Natalicchio, A, Laviola, L, De Tullio, C, Renna, LA, Montrone, C, Perrini, S, Valenti, G, Procino, G, Svelto, M and Giorgino, F (2004) Role of the p66Shc isoform in insulin-like growth factor I receptor signaling through MEK/Erk and regulation of actin cytoskeleton in rat myoblasts. J. Biol. Chem. 279:43900-9

To investigate the role of Shc in IGF action and signaling in skeletal muscle cells, Shc protein levels were reduced in rat L6 myoblasts by stably overexpressing a Shc cDNA fragment in antisense orientation (L6/Shcas). L6/Shcas myoblasts showed marked reduction of the p66Shc protein isoform and no change in p52Shc or p46Shc proteins compared with control myoblasts transfected with the empty vector (L6/Neo). When compared with control, L6/Shcas myoblasts demonstrated 3-fold increase in Erk-1/2 phosphorylation under basal conditions and blunted Erk-1/2 stimulation by insulin-like growth factor I (IGF-I), in the absence of changes in total Erk-1/2 protein levels. Increased basal Erk-1/2 activation was paralleled by a greater proportion of phosphorylated Erk-1/2 in the nucleus of L6/Shcas myoblasts in the absence of IGF-I stimulation. The reduction of p66Shc in L6/Shcas myoblasts resulted in marked phenotypic abnormalities, such as rounded cell shape and clustering in islets or finger-like structures, and was associated with impaired DNA synthesis in response to IGF-I and lack of terminal differentiation into myotubes. In addition, L6/Shcas myoblasts were characterized by complete disruption of actin filaments and cell cytoskeleton. Treatment of L6/Shcas myoblasts with the MEK inhibitor PD98059 reduced the abnormal increase in Erk-1/2 activation to control levels and restored the actin cytoskeleton, re-establishing the normal cell morphology. Thus, the p66Shc isoform exerts an inhibitory effect on the mitogen-activated protein kinase signaling pathway in rodent myoblasts, which is necessary for maintenance of IGF responsiveness of the MEK/Erk pathway and normal cell phenotype.

PubMed Online version:10.1074/jbc.M403936200

Adaptor Proteins, Signal Transducing/deficiency; Adaptor Proteins, Signal Transducing/genetics; Adaptor Proteins, Signal Transducing/physiology; Animals; Antibodies, Monoclonal; Base Sequence; Cells, Cultured; DNA Primers; Enzyme Inhibitors/pharmacology; Flavonoids/pharmacology; Mitogen-Activated Protein Kinase 1/metabolism; Mitogen-Activated Protein Kinase 3/metabolism; Myoblasts, Skeletal/cytology; Myoblasts, Skeletal/physiology; Phosphorylation; Polymerase Chain Reaction; Protein Subunits/immunology; Rats; Receptor, IGF Type 1/immunology; Receptor, IGF Type 1/physiology; Shc Signaling Adaptor Proteins; Signal Transduction; Transfection