Dissecting the role and pathogen benefits of trypanothione synthetase overexpression in Trypanosoma cruzi

ANDREA C. MESIAS; NATALIA SASONI; DIEGO G. ARIAS; NISHA J. GARG; MARÍA P. ZAGO

Baltimore

Congreso; American Society of Tropical Medicine and Hygiene Annual Meeting; 2017

American Society of Tropical Medicine and Hygiene

Chagas disease is atropical neglected illness caused by Trypanosomacruzi that remains to be endemic in Latin America. This pathogen has todeal with different oxidant challenges such as the respiratory burst triggeredinside the macrophage. Thus, it has an effective antioxidant system capable of overcomingthe host barriers and maintaining the redox balance. The parasite antioxidantnetwork utilizes trypanothione (TSH), a low MW dithiol, as substrate. The trypanothionesynthetase (TryS) enzyme (produces TSH metabolite) is uniquely present in kinetoplastids,and so it is a good candidate for drug design. In order to characterize itsrole in the host-parasite interaction, we have overexpressed pTREX encoding TrySin T. cruzi SylvioX10 isolate byelectroporation. Recombinant parasites (TryS+) exhibited stableoverexpression (>2-fold increase) of the TryS protein and a significantincrease in TryS enzymatic activity as compared to controls. The TryS+showed a higher rate of metacyclogenesis, and ~20% more of infective forms wereobtained in TryS+ cultures compared to the cultures of parasitestransfected with empty pTREX. Furthermore, transfectant parasites tolerated higherdoses of benznidazole (IC50 value: 21.3 µM and 11 µM, TryS+vs. controls, respectively, p<0.05). To get insight into TryS role in host-parasiteinteraction, HL-1 cells were infected. After 36 h incubation, cells infectedwith TryS+ (vs. control) T.cruzi exhibited 2-fold lower levels of intracellular ROS, determined by H2DCFDAfluorescence (p<0.05).Our results suggest that T. cruziutilizes TryS to promote differentiation from insect stage to infective formsand to maintain the redox balance in the host cells that support intracellularsurvival of the parasite.