Identifying the Role of EMX2 and the Network of EMX2‐sensitive Genes in Cell Lineage‐dependent Muscle Fiber Type Specification

Abstract

The mechanism of intrinsic muscle fiber diversification during development remains unclear. Slow myosin heavy chain 2 (MyHC2) gene expression designates fast/slow from fast muscle fibers in avian development. In this study, we examined the mechanism of regulation of lineage-specific fiber type development by analysis of slow MyHC2 promoter activity. EMX2 was identified by microarray analysis as a transcription factor that is differentially expressed in myoblasts and muscle fibers of the fast/slow versus fast myogenic cell lineage. The effect of EMX2 gene expression in primary muscle fibers was assessed by EMX2 gene overexpression and knockdown. These findings reveal a role for EMX2 transcription factor-dependent fiber type lineage specification. As cell lineage dependent myogenesis is a concerted effort by the contribution of multiple regulatory pathways, we aimed to identify EMX2-dependent genes within networks that define fast/slow versus fast myoblast lineage specification of distinct primary muscle fiber types. A genome-wide expression analysis in fast/slow myogenic cell clones with stably incorporated EMX2 was performed using RNA-Seq and was coupled to bioinformatics to identify genes differentially regulated with the overexpression of EMX2. Here we present preliminary verification of EMX2-associated factors identified by our RNA-Seq study, which are hypothesized to play a role within slow MyHC2 gene regulation. Identification and elucidation of this network of transcriptional regulators will give further insight into the greater network of gene interactions in skeletal muscle fiber phenotype specification during development.