EIF3 is the largest multi-protein complex, and several studies have revealed the oncogenic roles of its subunits in many human cancers. However, the roles of EIF3D in the development and progression of PCa remain uncovered. In the present study, the expression of EIF3D in prostate cancer and paracarcinoma tissues, as well as PCa cell lines, was examined. In PCa tissues, the expression of EIF3D was up-regulated compared to that in paracarcinoma tissues. In order to investigate whether EIF3D could serve as potential therapeutic target for prostate cancer, EIF3D was knocked down to verify its functional role in prostate cancer cells. After EIF3D knockdown in PC-3 and DU145 cells, cell proliferation, invasion and colony formation were significantly inhibited; meanwhile, cell cycle analysis revealed cell cycle arrest at G2/M phase. EIF3D is associated with PCa, and silencing EIF3D will result in decreased proliferation, and migration, as well as G2/M arrest in DU145 and PC-3 cells. These results suggest that EIF3D plays an oncogenic role in PCa development and progression.