UCLA

Toxoplasma gondii is an obligate intracellular parasite that can infect any warm-blooded mammal, making it one of the most successful parasites in the world. It is estimated to infect approximately one-third of the global human population and can cause life threatening complications in immunocompromised individuals and congenitally infected neonates. During the acute infection, rapidly multiplying T. gondii tachyzoites must balance between avoiding clearance to ensure the infection and limiting parasite replication to avoid killing the host. While most tachyzoites are eliminated during the acute infection by the host’s immune response, a fraction converts into slow growing bradyzoites that are encased by a cyst wall that constitute the chronic infection. These infectious tissue cysts remain largely undetected from the host’s immune surveillance.The ability of T. gondii to survive intracellularly and establish a productive infection relies significantly on its capacity to regulate host cell functions. One way the parasite achieves this is through the secretion of dense granule proteins (GRAs) that are involved in remodeling the parasitophorous vacuole (PV), obtaining nutrients from the host, manipulating the host cell cycle, and modulating the immune response. Because of their diverse functions during infection, the goal of this work is to identify and characterize novel GRAs in the acute and chronic infection to better understand the array of effectors this parasite uses to colonize its mammalian hosts. This work first describes a proximity labeling experiment to identify several novel T. gondii GRAs, GRA55-59, that are expressed in bradyzoites. Deletion of these GRAs does not affect parasite fitness in vitro. Further characterization of GRA55 reveals that its deletion does not impact virulence during the acute infection but does reduce brain cyst burden in infected mice. This suggests that GRA55 is an important secreted protein for the establishment or maintenance of the chronic infection in mice. The next section identifies the dense granule protein GRA83 and reveals that this effector is secreted into the PV and plays a role in modulating a key element of the host’s innate immune response. Disruption of GRA83 impacts proinflammatory cytokine production and increases T. gondii’s virulence during the acute infection. This also results in a significantly higher brain cyst burden during the chronic infection in mice. Therefore, this effector functions in a pro-host manner to limit unrestricted parasite growth. Lastly, this work describes the novel secreted effector, GRA84, that is exported to the host cell nucleus. GRA84 is dependent on the aspartyl protease (ASP5) and the MYR translocon to traverse the PV barrier and reach the host cell nucleus. Disruption of GRA84 does not substantially impact in vitro parasite growth or the chronic infection in mice. Most interestingly, this work demonstrates that GRA84 is proteolytically cleaved for maturation in its N-terminus and that this processing event is essential its ability to translocate across the vacuolar membrane to reach its destination in the host nucleus. Together, this thesis identifies and characterizes an array of novel GRA proteins that regulate Toxoplasma infection, which is likely to aid in the development of new treatments or therapeutics that can ultimately clear this widespread human infection.