AIM To determine the role of G0/G1 switch gene 2(G0 S2) and its transcriptional regulation in palmitate-induced hepatic lipid accumulation.METHODS Hep G2 cells were treated with palmitate,or palmitate in combination with CCAAT/enhancer binding protein(C/EBP)β si RNA or G0 S2 si RNA. The m RNA expression of C/EBPβ,peroxisome proliferator-activated receptor(PPAR)γ and PPARγ target genes(G0 S2,GPR81,GPR109 A and Adipoq) was examined by q PCR. The protein expression of C/EBPβ,PPARγ,and G0 S2 was determined by Western blotting. Lipid accumulation was detected with Oil Red O staining and quantified by absorbance value of the extracted Oil Red O dye. Lipolysis was evaluated by measuring the amount of glycerol released into the medium. RESULTS Palmitate caused a dose-dependent increase in lipid accumulation and a dose-dependent decrease in lipolysis in Hep G2 cells. In addition,palmitate increased the m RNA expression of C/EBPβ,PPARγ,and PPARγ target genes(G0 S2,GPR81,GPR109 A,and Adipoq) and the protein expression of C/EBPβ,PPARγ,and G0 S2 in a dose-dependent manner. Knockdown of C/EBPβ decreased palmitate-induced PPARγ and its target genes(G0 S2,GPR81,GPR109 A,and Adipoq) m RNA expression and palmitate-induced PPARγ and G0 S2 protein expression in Hep G2 cells. Knockdown of C/EBPβ also attenuated lipid accumulation and augmented lipolysis in palmitate-treated Hep G2 cells. G0 S2 knockdown attenuated lipid accumulation and augmented lipolysis,while G0 S2 knockdown had no effects on the m RNA expression of C/EBPβ,PPARγ,and PPARγ target genes(GPR81,GPR109 A and Adipoq) in palmitate-treated Hep G2 cells. CONCLUSION Palmitate can induce lipid accumulation in Hep G2 cells by activating C/EBPβ-mediated G0 S2 expression.