Translationally controlled tumor protein (TCTP) has various cellular functions and molecular interactions, many related to its growth-promoting and antiapoptotic properties. Recently, TCTP expression was reported to increases in insulin-resistant mice fed with high-fat diet. TCTP is a multifunctional protein, but its role in liver metabolism is unclear. Here, we investigated the function and mechanism of TCTP in HepG2 cells. Knock-down of TCTP led to 287 differentially expressed genes (DEGs) that were highly associated with cellular apoptosis and signal response, TNF and NF-kappa B signaling pathways, glycolysis/gluconeogenesis, insulin resistance, FoxO and insulin signaling pathways, adipocytokine and AMPK signaling pathways. shTCTP downregulated the expression of the key gluconeogenesis enzyme phosphoenolpyruvate carboxykinase (PCK1). Furthermore, TCTP regulated the alternative splicing of genes enriched in the phospholipid biosynthetic process and glycerophospholipid metabolism. We further showed that shTCTP down-regulated the intracellular levels of triglyceride and total cholesterol. Our results showed that TCTP regulates the liver cell transcriptome at both the transcriptional and alternative splicing levels. The TCTP regulatory network predicts the biological functions of TCTP in glucose and lipid metabolism, and also insulin resistance, which may be associated with liver metabolism and diseases such as nonalcoholic fatty liver disease.