Loss of Seipin causes the absence of whole-body adipose but abnormal liver lipid deposition in patients, and liver expression of Seipin is decreased in mice fed a high-fat diet (HFD). However, the underlying mechanism of Seipin-regulated liver lipid metabolism remains mysterious. Here, experiments show that over-expression of Seipin down-regulates HFD-induced liver triglyceride (TG) accumulation and promotes zebrafish growth. Real-time PCR and immunoblotting suggest that Seipin interacts with Plin2 through its second transmembrane domain to inhibit the expression of Plin2 by promoting Plin2 ubiquitination, thereby ameliorating lipid accumulation. Co-immunoprecipitation (CoIP) experiments and Biomolecular fluorescence complementation (BiFC) analysis reveal a close interaction between Seipin and phosphatidylcholine (PC) synthesis enzyme CCTα and CHPT in zebrafish and mice. Thus, Seipin may participate in PC synthesis and increase cellular PC levels. Elevated PC levels subsequently suppress Plin2 expression. Meanwhile, CCTα, the rate-limiting enzyme of the PC synthesis pathway, exhibited a unique regulatory role on Plin2 expression as a potential transcription factor. It is proposed that Seipin balances TG homeostasis, PC synthesis, and Plin2 expression to alleviate hepatic steatosis, providing a promising target for fatty liver disease.