Abstract
The obligate intracellular protozoan Toxoplasma gondii resides within a specialized parasitophorous vacuole (PV), isolated from host vesicular traffic. In this study, the origin of parasite cholesterol was investigated. T. gondii cannot synthesize sterols via the mevalonate pathway. Host cholesterol biosynthesis remains unchanged after infection and a blockade in host de novo sterol biosynthesis does not affect parasite growth. However, simultaneous limitation of exogenous and endogenous sources of cholesterol from the host cell strongly reduces parasite replication and parasite growth is stimulated by exogenously supplied cholesterol. Intracellular parasites acquire host cholesterol that is endocytosed by the low-density lipoprotein (LDL) pathway, a process that is specifically increased in infected cells. Interference with LDL endocytosis, with lysosomal degradation of LDL, or with cholesterol translocation from lysosomes blocks cholesterol delivery to the PV and significantly reduces parasite replication. Similarly, incubation of T. gondii in mutant cells defective in mobilization of cholesterol from lysosomes leads to a decrease of parasite cholesterol content and proliferation. This cholesterol trafficking to the PV is independent of the pathways involving the host Golgi or endoplasmic reticulum. Despite being segregated from the endocytic machinery of the host cell, the T. gondii vacuole actively accumulates LDL-derived cholesterol that has transited through host lysosomes.
Highlights
Upon entering a host cell, many intracellular pathogens reside within membrane-bound vacuoles
We have addressed the following questions, relating to the origin of cholesterol for the parasite: Can T. gondii synthesize its own cholesterol via the classical mevalonate pathway? Is the parasitophorous vacuole (PV) accessible to host cell cholesterol? If accessible, is it the cholesterol synthesized by the host cell or the exogenous cholesterol delivered by low-density lipoprotein (LDL) endocytosis that can be transported into the parasite? If acquired exogenously from LDL, is cholesterol transported from lysosomes to the PV by a direct transfer, a Golgi, or an ER-dependent pathway? Is the host cell altered in its cholesterol biosynthesis or LDL uptake in response to parasitization? Is the parasite capable of replication in host cells unable either to synthesize cholesterol de novo, or to use LDL-delivered cholesterol, or both?
The Toxoplasma PV remains segregated from vesicular trafficking through the endo- and exocytotic pathways in the host cell, the results presented here demonstrate that this parasite can actively intercept host LDLderived cholesterol, in transit from the lysosomes to other cellular compartments
Summary
Upon entering a host cell, many intracellular pathogens reside within membrane-bound vacuoles. Tally isolated from the host cell vesicular transport system (Jones et al, 1972; Sibley et al, 1985; Joiner et al, 1990; Mordue et al, 1999) This parasite is auxotrophic for several metabolites (see review by Sinai and Joiner, 1997) and must exchange nutrients across the PV membrane (PVM), surrounding it to assure its survival and propagation. This raises the intriguing issue of how nutrients are obtained from the host cell by T. gondii. Within the vacuolar space is a tubulo-reticular network connected to the PVM, which likely increases the exchange surface between the host cytoplasm and intravacuolar parasites (Sibley et al, 1995)
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