Gain-of-function PfAAT1 mutations compensate for impaired PfCRT function in Plasmodium falciparum

Area of Science:

• Molecular parasitology
• Drug resistance mechanisms
• Population genetics

Background:

• Mutations in the chloroquine resistance transporter (PfCRT) of *P. falciparum* confer resistance to chloroquine (CQ).
• Mutant PfCRT impairs the export of hemoglobin-derived peptides from the parasite digestive vacuole.
• PfAAT1 mutations are linked to CQ resistance, but the mechanism is unclear.

Purpose of the Study:

• Investigate the functional relationship between PfCRT and PfAAT1.
• Elucidate the mechanistic basis of PfAAT1's role in CQ resistance.
• Determine the evolutionary drivers of PfAAT1 mutations.

Main Methods:

• Isogenic parasite lines with wild-type and mutant *pfcrt* alleles.
• Knockdown of PfAAT1 expression.
• Analysis of hemoglobin-derived peptide accumulation.
• Comparative growth assays under nutrient-limiting conditions.

Main Results:

• PfAAT1 is essential in parasites with mutant PfCRT but dispensable with wild-type PfCRT.
• PfAAT1 knockdown leads to hemoglobin-derived peptide accumulation, indicating functional overlap with PfCRT.
• Field-derived PfAAT1 variants (S258L, F313S) provide a fitness advantage in mutant PfCRT backgrounds.
• The F313S mutation confers a fitness advantage under cysteine- and methionine-limiting conditions.

Conclusions:

• PfAAT1 compensates for impaired PfCRT function in malaria parasites.
• Selection of *pfaat1* alleles is driven by epistasis with *pfcrt* mutations.
• PfAAT1 mutations do not independently affect CQ sensitivity.