ZBED6 Knockout Promotes Skeletal Muscle Development in Bama Pigs by Relieving Transcriptional Repression of CDKN1A

Skeletal muscle development not only determines the growth performance and meat quality of livestock, but is also closely associated with human metabolic health. ZBED6 has been identified as a transcriptional repressor of IGF2 , yet its role and molecular mechanism in skeletal muscle development remain unclear. In this study, we employed a ZBED6 knockout (KO) Bama pig model and found that loss of ZBED6 led to increased muscle mass and muscle fiber hypertrophy, suggesting that ZBED6 plays an inhibitory role in muscle growth. To further elucidate the underlying mechanisms, we performed integrated transcriptomic and functional analyses to investigate the ZBED6–CDKN1A axis. RNA sequencing identified differentially expressed genes across seven tissues from 8‐month‐old KO and wild‐type pigs, and WGCNA revealed a skeletal muscle‐specific module enriched in myogenic pathways. By integrating this module with transcriptome data from longissimus dorsi at 5 and 8 months of age and ChIP‐seq data, CDKN1A was identified as a central hub gene. Dual‐luciferase reporter assays demonstrated that ZBED6 binds to a conserved GCTCG motif in the CDKN1A promoters of pigs and mice to repress its transcription. Consistently, in C2C12 cells, ZBED6 knockdown upregulated the expression of myogenic markers ( MyoD , MyoG , and MyHC ) and promoted myotube formation, whereas silencing CDKN1A attenuated this effect. Conversely, ZBED6 overexpression reduced CDKN1A expression and suppressed differentiation, confirming its bidirectional regulatory role. Collectively, our data identify CDKN1A as a transcriptional target of ZBED6 and show that ZBED6 constrains skeletal muscle development by repressing CDKN1A .