QKI Promotes Sheep Preadipocyte Differentiation by Reducing Cavin3 mRNA Stability

Adipogenic differentiation is essential for adipose tissue development, fat deposition, and metabolic regulation in livestock, but the role of RNA-binding proteins in sheep preadipocyte differentiation remains unclear. This study investigated whether Quaking (QKI) regulates sheep preadipocyte adipogenesis and explored its downstream regulatory mechanism. QKI expression was examined in sheep adipose tissues, and QKI knockdown was performed in sheep preadipocytes. Adipogenic marker expression, lipid accumulation, glucose consumption, Oxford Nanopore Technologies (ONT) full-length transcriptome sequencing, RNA immunoprecipitation coupled with quantitative real-time PCR (RIP-qPCR), actinomycin D assay, caveolae-associated protein 3 (Cavin3) knockdown, and simultaneous QKI and Cavin3 knockdown were used to evaluate the function and mechanism of QKI. QKI was expressed in different sheep adipose tissues, with relatively higher expression in tail fat. QKI knockdown reduced adipogenic marker expression, Oil Red O staining, and glucose consumption after adipogenic induction. ONT full-length transcriptome sequencing identified Cavin3 as a markedly upregulated candidate downstream target after QKI knockdown. RIP-qPCR showed enrichment of Cavin3 mRNA in QKI immunoprecipitates, and actinomycin D assays indicated that QKI knockdown delayed Cavin3 mRNA degradation. Cavin3 knockdown enhanced adipogenic marker expression, Oil Red O staining, and glucose consumption. Simultaneous QKI and Cavin3 knockdown increased adiponectin, fatty-acid-binding protein 4, and peroxisome proliferator-activated receptor gamma mRNA abundance and Oil Red O staining relative to the matched control, whereas glucose consumption was not significantly different. These findings suggest that QKI promotes sheep preadipocyte differentiation, at least in part, by reducing Cavin3 mRNA stability, providing insight into RNA-binding protein-mediated regulation of livestock adipogenesis.