Heme serves as a crucial cofactor for numerous enzymes, particularly those in the mitochondrial respiratory chain. Toxoplasma gondii can synthesize heme de novo through a biosynthesis pathway spanning the mitochondrion, apicoplast, and cytosol. Interestingly, the parasite can bypass heme synthesis by salvaging host cell heme intermediates, such as protoporphyrin IX (PPIX), which are elevated with excess 5-aminolevulinic acid (5-ALA)¹. However, the mechanisms and transporters facilitating the parasite's access to PPIX or other heme intermediates remain elusive.

To identify these transporters, we conducted genome-wide CRISPR-based forward genetic screens, comparing wildtype and mutant parasites with defects in the cytosolic enzyme coproporphyrinogen III oxidase (CPO). Mutants were rescued by the addition of 5-ALA, facilitating the identification of transporters involved in heme intermediate salvage. While these transporters are non-essential in wildtype parasites, they become indispensable in heme synthesis-deficient mutants. Three ABC transporters identified from the screen are predicted to localize to the plasma membrane and mitochondrion.

Our current efforts are focused on characterizing these candidate transporters to validate their localization and function. Given the limited understanding of metabolic needs during the chronic stage, we plan to employ an in vitro model system based on human myotubes capable of harboring mature cysts². Through this model, we aim to evaluate the significance and function of the newly identified transporters during the bradyzoite stage.

¹Krishnan A et al, Cell Host Microbe. 2020 Feb 12;27(2):290-306.e11.

²Christiansen C et al, Nat Commun. 2022 Mar 4;13(1):1168.