PLIN2 Protein Promotes Lipid Droplet Fusion in Ningxiang Pig (Sus scrofa domesticus) Through CIDea Protein
LI Ji-Hao1,3, YUE Chao2, CHEN Shu-Xun1, GU Hao3,*, MA Na2,*
1 College of Life Sciences, Nanyang Normal University, Nanyang 473061, China; 2 College of Water Resources and Modern Agriculture, Nanyang Normal University, Nanyang 473061, China; 3 Hubei Provincial Key Laboratory of Animal Embryo Engineering and Molecular Breeding, Institute of Animal Husbandry and Veterinary Medicine, Hubei Academy of Agricultural Sciences, Wuhan 430064,China
Abstract：The Ningxiang pig (Sus scrofa domesticus), a geographically indicated agricultural product of China, is characterized by excessive backfat thickness due to oversized lipid droplets, which represents a major disadvantage. To investigate the regulatory mechanisms of lipid droplet size and develop a more market-competitive strain of Ningxiang pigs, this study utilized PLIN2 (perilipin-2) knockout Ningxiang pigs as experimental models. Transmission electron microscopy was employed to observe lipid droplet size and fusion events, while qRT-PCR and Western blot were used to verify the expression of downstream target genes. Additionally, lipid droplet size was assessed using staining techniques such as boron-dipyrromethene (BODIPY) and Oil Red O. Following gene expression screening via qRT-PCR and validation through Western blot, CIDEa (cell death-inducing DNA fragmentation factor alpha-like effector A) was identified as a key regulator of lipid droplet size. A CIDEa overexpression vector was constructed and transfected into PLIN2-knockout Ningxiang pig cells, and lipid droplet size was subsequently evaluated. Furthermore, bioinformatics tools were utilized to predict the binding sites between PLIN2 and CIDEA. The results showed that PLIN2 knockout significantly reduced lipid droplet fusion and markedly decreased CIDEA expression. Overexpression of CIDEa compensated for the lipid droplet reduction caused by PLIN2 deletion. AlphaFold predictions revealed multiple binding sites between PLIN2 and CIDEa. These findings suggest that PLIN2 may regulate lipid droplet size through CIDEA-mediated lipid droplet fusion, providing novel theoretical insights into the regulatory mechanisms of lipid droplet size and a scientific basis for breeding more market-competitive Ningxiang pig strains.
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