New gene targets of PGC-1α and ERRa co-regulation in C2C12 myotubes

As a transcriptional coactivator, PGC-1 alpha contributes to the regulation of a broad range of metabolic processes in skeletal muscle health and disease; however, there is limited information about the genes it transcriptionally regulates. To identify new potential gene targets of PGC-la regulation, mouse C2C12 myotubes were screened by microarray analysis following PGC-1 alpha overexpression. Genes with an mRNA expression of 2.5-fold or more (P <0.001) were identified. From these, further genes were singled out if they had no previous connection to PGC-1 alpha regulation or characterization in skeletal muscle, or were unannotated with no known function. Following confirmation of their regulation by PGC-1 alpha using qPCR analysis, eight genes were focused on for further investigation (Akr1b10, Rtnndl, 1110008P14Rik, 1700021F05Rik, Mtfpl, Mrtnl, Oxnadl and Cluh). Bioinformatics indicated a number of the genes were linked to a range of metabolic-related functions including fatty acid oxidation, oxido-reductase activity, and mitochondrial remodeling and transport. Treating C2C12 myotubes for 6 h with AICAR, a known activator of AMP kinase and inducer of Pgc-1 alpha gene expression, increased the mRNA levels of both Pgc-1 alpha (P <0.001) and of Mtfpl, Mrtnl, Oxnadl and Cluh (P < 0.05). Screening of the promoter and intron 1 regions also revealed all genes to contain either a consensus or near consensus response elements for the estrogen-related receptor a (ERR alpha), a key transcription factor-binding partner of PGC-1 alpha in skeletal muscle. Furthermore, knockdown of endogenous ERRa levels partially or completely blocked the induction of gene expression of all genes by PGC-1 alpha, while each gene was significantly upregulated in the presence of a constitutively active form of ERRa (P < 0.05) except for Akr1b10. These findings provide preliminary evidence for the novel regulation of these genes by PGC-1 alpha and its signaling pathway in skeletal muscle.