Authors

Discussion

Trypanosoma cruzi, the causing agent of Chagas’ disease, is highly adapted to life in diverse environments, and part of this adaptation is conferred by the metabolic flexibility these organisms have. In this context, the metabolism of amino acids plays different roles in biological processes that allow T. cruzi to survive, replicate, and progress through its life cycle. Our lab has previously characterised the ability of T. cruzi epimastigotes to uptake and oxidize histidine (His)which points to the existence of an active His degradation pathway. In other organisms, this pathway comprises four enzymatic steps, being the first the non-oxidative deamination of His to urocanate, catalysed by histidine ammonia-lyase (HAL). To investigate the biological roles of the putative TcHAL, we generated T. cruzi CL Brener epimastigotes TcHAL null mutants using CRISPR-Cas9. TcHAL null mutants showed no proliferation phenotype when compared to the wild type and Cas9-expressing controls. When TcHAL null parasites were incubated with [¹⁴C(U)]-His, they did not produce considerable levels of radiolabelled CO_2, showing the successful disruption of the His degradation pathway. When TcHAL null mutants were recovered from nutritional stress with His, they were not able to use His to trigger oxygen consumption. Interestingly, high concentrations of His were toxic to TcHAL null mutants in nutritional stress. Finally, the double knockout of TcHAL affected the capability of these parasites to differentiate in vitro in the presence of different substrates but did not alter the infection rate on the triatomine vector Rhodnius prolixus when compared to the Cas9-expressing control. Together, our data show that disruption of the putative TcHAL coding sequence leads to inactivation of the first step of His degradation pathway affecting the ability of the parasite to metabolise and use His for mitochondrial ATP synthesis, but with no consequences to the infection of the insect vector.