Interplay of Trypanosoma cruzi and host metabolism and impact on parasite intracellular growth

Abstract

The protozoan parasite Trypanosoma cruzi, which causes Chagas disease, is capable of infecting and replicating within metabolically distinct tissues such as the heart, skeletal muscle, and adipose tissue, driving disease pathogenesis. However, nutrient requirements of the replicative intracellular amastigote form and knowledge of how host metabolism affects parasite growth remain ill defined. Studying the metabolism of intracellular amastigotes has been difficult given the overlap between parasite and mammalian metabolic pathways and the challenges of obtaining enough amastigote material for biochemical characterization, necessitating the development of new methodologies. In this dissertation we study amastigote and host metabolism during T. cruzi infection using novel techniques. Predictions that amastigotes oxidize fatty acids for energy generation were investigated using microscopy and extracellular flux analysis, which revealed that intracellular amastigotes take up a form of exogenous palmitate, likely palmitoyl-CoA, which stimulates amastigote mitochondrial respiration. Knockdown of host CPT1, to reduce entry of cytosolic fatty acyl-CoA into mitochondria for beta-oxidation, increased parasite intracellular growth, pointing to a role for host cytosolic long chain fatty acyl-CoA pools in sustaining intracellular parasite replication. To characterize how T. cruzi metabolically adapts to intracellular infection, we paired transcriptome data with functional data from an optimized mitochondrial stress test for different life cycle stages. Transcriptome and extracellular flux analysis revealed increased metabolic capabilities of intracellular amastigotes compared to extracellular trypomastigotes or insect stage epimastigotes. However, amastigote growth in different host metabolic states did not induce transcriptional metabolic reprogramming or substantial alterations to the bioenergetic profile, suggesting that increased amastigote metabolic plasticity aids adaptation to multiple different intracellular milieus. To determine whether T. cruzi infection also modulates host metabolism to improve intracellular infection, we examined the impact of infection on host mitochondrial respiration and glucose uptake. Inhibition of amastigote mitochondrial respiration in infected monolayers with ELQ300 revealed that host cells increase mitochondrial respiration during infection, though that increase is dispensable for parasite replication. However, labeling assays demonstrated that infection drives increased glucose uptake in the host, and the amastigote accesses host cytosolic glucose which it uses to support energy generation, biosynthetic processes, and replication. In summary, the data indicate that amastigotes increase their metabolic capacity and modulate host metabolism to increase access to key nutrients, which fuels intracellular replication.