In vitro overexpression of CHAPa and CHAPb in mouse cardiomyocytes and skeletal muscle cells interferes with Z-disc integrity and decreases fetal gene expression

In the previous chapter we have described that CHAPb transgenic mice displayed features of cardiomyopathy and cardiac dysfunction, whereas CHAPa transgenic mice did not display any abnormalities. In order to determine direct e ects of both CHAP isoforms, we infected mouse cardiomyocytes (E17.5) and skeletal muscle cells myoblast cells (C2C12) with adenoviral constructs for CHAPa and CHAPb. In vitro overexpression of CHAPa resulted in Z-disc disruption, as shown by α-actinin-2 staining in both C2C12 cells and cardiomyocytes. CHAPb overexpression in C2C12 and cardiomyocytes resulted in stress ber formation, which stained for F-actin. Although RhoA and phosphorylated Ezrin/Moesin/Radixin, proteins involved in linking the actin cytoskeleton to the plasma membrane, were ectopically expressed in cardiomyocytes, no activation of the actin signaling pathway was observed. Surprisingly, in both CHAPa and CHAPb infected cells downregulation of hypertrophy markers Nppa, Nppb and Myh7 was observed, whereas Myh6 was upregulated. In agreement with these ndings we observed that a key player in muscle hypertrophy and development, the transcription factor Nuclear Factor of Activated T-cells-c2 (NFATc2), was translocated from the nucleus to the cytoplasm. ese results show that CHAPa and CHAPb have an important role in maintaining the integrity of muscle cells. Furthermore, interestingly our ndings suggest that CHAPa and b isoforms may directly a ect transcriptional regulation regarding maturation and/or blocking hypertrophy of cardiomyocytes.