Although parasite-infected host cells become resistant to apoptosis, uninfected bystander cells undergo apoptosis during Toxoplasma gondii infection. The Programmed Cell Death 5 (TgPDCD5) gene, a homologue of the human apoptosis-related molecule, was cloned from a T. gondii full-length cDNA database and subsequently characterized. The native TgPDCD5 was located in the cytosol and also detected in the secreted fraction. Immuno-electron microscopic analysis showed TgPDCD5 was primarily located close to the rhoptries or vesicle-like structures near the surface membrane of the parasite. Studies using recombinant TgPDCD5 (rTgPDCD5) demonstrated that host cells internalize the molecule in a heparan sulfate proteoglycan-binding motif-dependent manner. Furthermore, the addition of rTgPDCD5 to culture medium resulted in the enhancement of host-cell apoptosis triggered by etoposide in macrophage cell line J774A.1 and leukemic cell line HL-60 cells. Additionally, rTgPDCD5 induced apoptosis in J774A.1 cells in the presence of IFN-gamma. This report is the first to identify a parasitic molecule of T. gondii that has a pro-apoptotic effect on host cells.