Myostatin (MSTN), associated with t

Myostatin (MSTN), associated with the "double muscling" phenotype, affects muscle growth and fat deposition in animals. We aimed to uncover the molecular mechanism via which MSTN regulates adipogenesis. We generated MSTN-knockout (KO) cloned Meishan pigs using an improved gene targeting strategy relying on dual fluorescence-assisted selection (DUFAS). MSTN-KO pigs exhibits classical double muscling trait that features enhanced leanness and reduced fatness. We sequenced transcriptome of subcutaneous fat tissues of wild-type (WT) and MSTN-KO pigs, and intersected the differentially expressed mRNAs and miRNAs to predict that stearoyl-CoA desaturase 5 (SCD5) is targeted by miR222. Transcription factor binding prediction showed that myogenic transcription factor 2C (MEF2C) potentially binds to the miR222 promoter. We hypothesized that MSTN-KO upregulates MEF2C and consequently increases the miR222 expression, which in turn targets SCD5 to suppress its translation. Loss- and gain-of functional analysis of miR222 revealed that SCD5 was a bona fide target of miR222, which reduced nonsaturated palmitoleic (C16:1) and oleic (C18:1) acids and triglyceride (TG) synthesis. Overexpression and silencing of MEF2C revealed that miR222 upregulation by MSTN-KO was MEF2C-dependent. ChIP and EMSA analyses verified that MEF2C interacts with the miR222 promoter. Altogether, we report that MSTN affects adipogenesis through the MEF2C/miR222/SCD5 cascade.