We recently showed that the neurotropic parasite Toxoplasma gondii has a complete endolysosomal system, including a lysosome-like vacuolar compartment (VAC). Additional recent work showed that T. gondii uses its endolysosomal system to acquire proteins from the cytoplasm of infected cells, unexpectedly revealing that Toxoplasma is a heterophage (literally meaning other eat). Herein we show that disrupting VAC proteases also causes accumulation of autophagosomes in chronic stage Toxoplasma, suggesting that protease deficient parasites are resource limited and that the VAC contributes to digestion of autophagosomes. We also show that parasites deficient in VAC proteases perish in vitro and in vivo, creating an exciting opportunity to gain new insight into mechanisms of long-term parasite survival. Based on these findings we propose that parasites lacking VAC-resident proteases die because of a failure to obtain amino acids from degradation of host-derived proteins and autophagosomes. We will test this hypothesis by: (1) identifying the role of VAC-associated proteases in bradyzoite viability; (2) defining mechanisms of autophagy in protease deficient bradyzoites; and (3) determining the role of amino acid limitation in the demise of protease deficient bradyzoites. These studies move toward our long-term goal of understanding mechanisms of key processes required for T. gondii bradyzoite survival and chronic infection, which occurs in an estimated 2 billion people globally and 60 million Americans. HIV positive individuals are at risk of developing reactivated toxoplasmosis manifested as fatal encephalitis, myocarditis or loss of vision. The studies proposed herein will help reveal how T. gondii manages resources to sustain long-term survival during persistent infection. The absence of this knowledge precludes new opportunities to exploit deficiencies in parasite proteolytic digestion and potentially reduce the risk of reactivated toxoplasmosis.
We seek to understand how the brain parasite Toxoplasma gondii persists in infected individuals with the long-term goal of exploiting this knowledge for new treatments for AIDS-associated toxoplasmosis. This project will follow up on our recent finding that Toxoplasma appears to eat itself during chronic infection via a process called autophagy. Disruption of autophagy in parasites lacking certain proteases leads to parasite death. The work will reveal new knowledge of how Toxoplasma survives indefinitely in the brain, potentially leading to new opportunities to minimize the risk of reactivated disease in HIV positive individuals.
|Di Cristina, Manlio; Carruthers, Vern B (2018) New and emerging uses of CRISPR/Cas9 to genetically manipulate apicomplexan parasites. Parasitology 145:1119-1126|
|Di Cristina, Manlio; Dou, Zhicheng; Lunghi, Matteo et al. (2017) Toxoplasma depends on lysosomal consumption of autophagosomes for persistent infection. Nat Microbiol 2:17096|
|Guerra, Alfredo J; Carruthers, Vern B (2017) Structural Features of Apicomplexan Pore-Forming Proteins and Their Roles in Parasite Cell Traversal and Egress. Toxins (Basel) 9:|
|McGovern, Olivia L; Carruthers, Vern B (2016) Toxoplasma Retromer Is Here to Stay. Trends Parasitol 32:758-760|