Hemizygous mutation of Jmjd3 in muscle stem cells increases H3K27 methylation on Pax7 leading to impaired myogenesis.

Development of myogenic cells, called satellite cells, is determined by transcription factors that regulate their quiescence (e.g., Pax7), activation (e.g., MyoD), and terminal differentiation (e.g., myogenin). Demethylation of lysine 3 on histone 27 (H3K27) activates expression of Myod and Myog. In this investigation, we investigated the effects of a satellite cell-targeted, hemizygous mutation of the H3K27 demethylase Jmjd3 in healthy muscle. Using sequencing of chromatin fragments precipitated from Jmjd3 mutant and control satellite cells using anti-H3K27me3, we found that the Pax7 promoter was the only chromatin that experienced significantly increased H3K27 methylation in mutant cells. However, RNA sequencing showed that 143 genes were downregulated in mutant cells, including Myod, a direct target of Pax7, and at least 72 other genes that contained E-boxes targeted by MyoD. Gene ontology analysis showed enrichment of genes involved in myogenesis in the downregulated genes. Reduced expression of Pax7, Myod, and Myog was confirmed by quantitative PCR (qPCR), Western blots, and immunohistochemistry. Mutant muscles also had smaller diameter fibers, fewer myonuclei, and diminished myogenic cell fusion, indicating impaired growth and differentiation. These findings show that Jmjd3 affects demethylation of H3K27 at the Pax7 promoter, and increased H3K27 methylation reduces expression of Pax7 and its target genes, disrupting muscle growth.NEW & NOTEWORTHY This investigation reveals a new mechanism that regulates the development of muscle. Mutating Jmjd3 produced epigenetic modifications to the transcription factor Pax7, reducing its expression and impairing muscle growth.