D230025D16Rik: A Novel Regulator of Muscle Cell Differentiation

Abstract

RationaleLoss of skeletal muscle mass and function is a common clinical feature and poor prognostic indicator in various diseases. A promising therapeutic approach of regenerative medicine for repair of skeletal muscle is through the delivery of cellular therapies targeting regulators of muscle regeneration (myogenesis). However, not all the proteins involved in this process have been identified. To address the gap in knowledge of proteins involved in myogenesis, we recently identified a novel gene, D230025D16Rik, which has never been characterized within the context of skeletal muscle growth and repair.MethodsThe role of D230025D16Rik in a C2C12 murine model of myoblast differentiation was assessed by using an siRNA loss‐of‐function approach. Differentiation of C2C12 myoblasts into myotubes in cells with intact D230025D16Rik and those with D230025D16Rik knockdown (KD) was determined by various experimental techniques, such as RT‐qPCR, microarray, immunofluorescence staining, and immunoblotting.ResultsD230025D16Rik has a mainly nuclear localization in myoblasts, whereas in myotubes, D230025D16Rik has both a nuclear and cytosolic localization. D230025D16Rik expression level is induced at the onset of differentiation (i.e., Day 1) and steadily increases as myoblasts fuse into myotubes. In response to D230025D16Rik KD, the expression of transcription factors MyoD1 and Myogenin and muscle‐specific transcripts Myosin Heavy Chain (MyHC) and Muscle Creatine Kinase (MCK) were upregulated such that a significant acceleration of myogenesis was observed. This observation was further confirmed by an analysis of transcriptomes of control and D230025D16Rik KD myoblasts that revealed an upregulation of myogenesis‐related transcripts in cells with D230025D16Rik KD. Phenotypically, we observe hypertrophic myotube formation at days 0 and 1 in response to D230025D16Rik KD, as indicated by immunofluorescence staining for MyHC. Finally, the presence of nuclear aggregates on D230025D16Rik KD myotubes indicates that D230025D16Rik is essential for proper myotube formation.ConclusionAltogether, these findings highlight, for the first time, the potential regulatory functions of D230025D16Rik during myogenesis.