Ras regulation of skeletal muscle differentiation and gene expression

This chapter describes the Ras regulation of skeletal muscle differentiation and gene expression. The chapter describes the routine procedures for maintaining myogenic cell lines in undifferentiated and differentiated states. Methods for transient and stable transfection are presented in the chapter, along with the standard procedure for muscle-specific luciferase reporter gene assays and for the immunostaining of muscle-specific proteins in differentiated muscle cells. Ras proteins are essential components in the signal transduction pathways controlling cell proliferation, differentiation, and apoptosis. Constitutive activation of Ras and its downstream effectors is a molecular feature common to a large percentage of human tumors. Ras proteins inhibit the expression of the muscle regulatory factors [MRFs: members of the basic helix-loop-helix (bHLH) transcription factor family] in both primary myoblasts and in immortalized myoblast cell lines. Furthermore, in a manner that is not completely understood, activated Ras proteins block the ability of MRF transcription complexes to activate muscle-specific gene expression.