Mechanisms driving reactivated toxoplasmosis in HIV/AIDS co-infection

About This Grant

PROJECT SUMMARY Toxoplasma gondii is an intracellular protozoan parasite that poses a major threat to patients co-infected with HIV. The parasite is transmitted through multiple routes, producing high seroprevalence worldwide, including 30-50% of those infected with HIV. The replicative stage (tachyzoite) develops into a latent stage (bradyzoite) that persists as cysts in brain, heart, and skeletal muscle. As immunity wanes during AIDS, tissue cysts reconvert into replicating tachyzoites, which cause rapid damage to critical organs. Reactivated AIDS- toxoplasmosis is recurrent and chronic, leading to serious encephalitis, seizures, and myocarditis. The molecular mechanisms driving the reactivation of toxoplasmosis require an understanding of how bradyzoites convert into tachyzoites, which has remained completely uninvestigated despite its clinical importance in the context of AIDS. Our laboratory has identified translational control to be pivotal in parasite stage transitions, as it shifts which mRNA transcripts are favored for protein synthesis. Our investigation of translational control has found TgIF2α phosphorylation to be undetectable in unstressed tachyzoites but prominent in bradyzoites. Accordingly, inhibitors of TgIF2α phosphorylation block the reactivation of toxoplasmosis. These promising findings provide a unique opportunity to fill a glaring gap in our knowledge regarding the mechanisms of reactivated toxoplasmosis during AIDS. Our goal is to leverage our expertise in translational control to identify the factors enabling latent Toxoplasma cysts to reactivate in HIV/AIDS. We will accomplish this goal by determining how TgIF2α is dephosphorylated during reactivated toxoplasmosis (Aim 1) and by identifying preferentially translated mRNAs that are central for recrudescent infection (Aim 2). Aim 1 will use both targeted and unbiased approaches to maximize success: as it is highly likely that TgPP1 dephosphorylates TgIF2α based on what is known in other eukaryotes, we will conduct pulldowns during recrudescence to identify the regulatory proteins unique to its recruitment to phosphorylated TgIF2α. In case TgPP1 is not involved, our unbiased approach will identify the targets of TgIF2α dephosphorylation inhibitors that block reactivation of infection. Aim 2 will employ ribosomal profiling to identify mRNAs preferentially translated during reactivated toxoplasmosis. We will further determine the role of factors identified in each aim in reactivated toxoplasmosis by assaying the phenotype of genetic mutants made to lack the target. Our results will have a sustained high impact on the field, addressing mechanisms underlying one of the most urgent clinically relevant questions in AIDS-toxoplasmosis.