Abstract
Toxoplasma gondii is an obligate intracellular parasite that is able to infect virtually any nucleated cell of all warm-blooded animals. The host cell factors important for parasite attachment, invasion, and replication are poorly understood. We screened a siRNA library targeting 18,200 individual human genes in order to identify host proteins with a role in T. gondii growth. Our screen identified 19 genes whose inhibition by siRNA consistently and significantly lowered parasite replication. The gene ontology categories for those 19 genes represented a wide variety of functions with several genes implicated in regulation of the cell cycle, ion channels and receptors, G-protein coupled receptors, and cytoskeletal structure as well as genes involved in transcription, translation and protein degradation. Further investigation of 5 of the 19 genes demonstrated that the primary reason for the reduction in parasite growth was death of the host cell. Our results suggest that once T. gondii has invaded and established an infection, global changes in the host cell may be necessary to reduce parasite replication. While siRNA screens have been used, albeit rarely, in other parasite systems, this is the first report to describe a high-throughput siRNA screen for host proteins that affect T. gondii replication.
Highlights
Toxoplasma gondii is an obligate intracellular parasite with a complex life cycle that can include both sexual and asexual stages
Released tachyzoites repeat the cycle in subsequent cells until immune pressure and other poorly understood physiological cues trigger the parasite to switch to bradyzoites and establish a chronic infection
Cell Culture and Growth of Parasites T. gondii RHDHXGPRT [14] were maintained as tachyzoites by passage on monolayers of human foreskin fibroblasts (HFFs; ATCC)
Summary
Toxoplasma gondii is an obligate intracellular parasite with a complex life cycle that can include both sexual and asexual stages. While sexual replication occurs only in felines, the asexual cycle can occur in all warm-blooded animals, including humans. The asexual forms of T. gondii include tachyzoites, the fast growing form found primarily during acute infection, and bradyzoites, the encysted form responsible for maintaining a chronic infection. After ingestion of bradyzoite cysts, the parasite reverts to its tachyzoite form, which disseminates throughout the body. Tachyzoites actively invade host cells, forming a parasitophorous vacuole where the parasites rapidly replicate until the host cell lyses, releasing more tachyzoites. Released tachyzoites repeat the cycle in subsequent cells until immune pressure and other poorly understood physiological cues trigger the parasite to switch to bradyzoites and establish a chronic infection (reviewed in [1])
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