Prostaglandin E2 (PGE2) has been implicated in hepatocellular carcinoma cell invasion. Recently, it was reported that Y box-binding protein 1 (YB-1) is closely correlated with malignancy. This study was designed to examine the mechanisms by which PGE2 increases YB-1 expression and promotes HCC cell invasion. PGE2 greatly enhanced HCC cell invasion through upregulation of the YB-1 protein, and the EP1 receptor is mainly responsible for this regulation. Src and EGFR were both activated by PGE2, which in turn increased the phosphorylation levels of p44/42 MAPK. Src, EGFR and p44/42 MAPK were all involved in PGE2-induced YB-1 expression. Chemical inhibitors and RNAi analysis all confirmed the role of mTOR complex 1 in YB-1 expression induced by PGE2. Furthermore, YB-1 was able to regulate the expression of a series of EMT-associated genes, which indicated that YB-1 could have the potential to control the epithelial-mesenchymal transition process in HCC cells. These findings reveal that PGE2 upregulated YB-1 expression through the EP1/Src/EGFR/p44/42 MAPK/mTOR pathway, which greatly enhanced HCC cell invasion. This study for the first time describes the mechanisms through which PGE2 regulates YB-1 expression and promotes HCC cell invasion.