CRISPR-Mediated PABPC1 Upregulation via ELF5 Binding Disruption Enhances Milk Protein and Lipid Biosynthesis in Goat Mammary Epithelial Cells

Abstract Post-transcriptional control of milk protein and lipid production in goats remains poorly understood. Here we combined ONT full-length transcriptome sequencing with RNA pull-down coupled to LC–MS/MS to identify RNA-binding proteins associated with milk protein mRNAs, and uncovered poly(A)-binding protein cytoplasmic 1 (PABPC1) as a central regulator. PABPC1 bound the 3′UTRs of casein and lipid-related transcripts such as CSN2, CSN3, LALBA, and FASN, stabilizing these mRNAs and promoting their translation, while dose-dependently modulating alternative polyadenylation (APA) site selection to reshape 3′UTR isoform usage. ELF5 directly bound the PABPC1 promoter, repressing its transcription. CRISPR/Cas9 editing of this binding motif relieved this repression, elevating PABPC1 levels and increasing casein accumulation and triglyceride synthesis in goat mammary epithelial cells. Our findings define an ELF5–PABPC1–APA regulatory module and nominate a tractable target for molecular breeding to enhance goat milk protein and lipid content.