Expression of Gαz in C2C12 cells restrains myogenic differentiation

The recent identification of Gαz expression in C2C12 myoblasts and its demonstrated interaction with the transcription factor Eya2 inferred an unanticipated role of Gαz in muscle development. In the present study, endogenous Gαz mRNA and protein expressions in C2C12 cells increased upon commencement of myogenesis and peaked at around 4-6 days after induction but were undetectable in adult skeletal muscle. Surprisingly, stable expression of recombinant Gαz in C2C12 myoblasts strongly suppressed myotube formation upon serum deprivation, and the constitutively active mutant GαzQL exerted more pronounced effects. Transcriptional activities of reporter genes responsive to early (MyoD, MEF2 and myogenin) and late (muscle creatine kinase and myosin heavy chain) myogenic markers were reduced by transiently expressed GαzQL. Membrane attachment of Gαz was apparently required for the suppressive effects because a fatty acylation-deficient Gαz mutant could not inhibit myogenin expression. Introduction of siRNA against Gαz enhanced myogenin-driven luciferase activity and increased myosin heavy chain expression. Immunostaining of C2C12 cells over-expressing Gαz showed delayed nuclear expression of myogenin and severe myotube deformation. Gαz expression was accompanied by reduced levels of Rock2, RhoA and RhoGAP, enhanced expression of Rnd3, and a reduction of serum-responsive factor-driven reporter activity. These results support a novel role of Gαz in restraining myogenic differentiation through the disruption of Rho signaling.