ZBED6 deletion activates FNDC1-mediated PI3K/AKT and MAPK/ERK1/2 signaling pathways to promote myogenesis in pigs

Abstract

Myogenesis is a tightly regulated biological process that is essential for skeletal muscle growth, regeneration, and livestock production. Although the transcription factor ZBED6 is a key regulator of skeletal muscle development, the downstream molecular mechanisms through which it controls myogenesis remain incompletely understood. Here, using ZBED6-deficient pigs and primary porcine skeletal muscle satellite cells, we sought to identify novel downstream targets of ZBED6 and elucidate the molecular mechanisms underlying ZBED6-mediated regulation of myogenesis. By integrating transcriptomic, quantitative proteomic, and ChIP-seq datasets, we identified fibronectin type III domain-containing protein 1 (FNDC1), a muscle-derived secreted factor, as a previously unrecognized direct transcriptional target of ZBED6. ChIP-qPCR and dual-luciferase reporter assays showed that ZBED6 directly binds to the FNDC1 promoter and represses its transcription. Functional analyses in primary porcine skeletal muscle satellite cells showed that ZBED6 knockdown markedly promoted myoblast proliferation, differentiation, and myotube formation, whereas FNDC1 knockdown partially attenuated these effects. Conversely, FNDC1 overexpression partially rescued the impaired myogenesis caused by ZBED6 overexpression. Mechanistically, ZBED6 deficiency promoted porcine myoblast proliferation and differentiation, at least in part, through FNDC1-mediated activation of the PI3K/AKT and MAPK/ERK1/2 signaling pathways. Consistently, pharmacological inhibition of either pathway significantly attenuated the enhanced porcine myoblast proliferation and differentiation induced by ZBED6 deficiency, supporting that activation of these signaling pathways contributes in part to the pro-myogenic effects mediated by the ZBED6-FNDC1 regulatory axis in porcine myoblasts. This study identifies FNDC1 as a novel direct transcriptional target of ZBED6, supporting the ZBED6-FNDC1 axis as a regulatory mechanism contributing to porcine myoblast proliferation and differentiation. These findings expand the current understanding of the ZBED6 regulatory network and provide a cellular framework for future studies investigating skeletal muscle growth in vivo.