FIG 7.

PfPX1 knockout lines are more sensitive to PI3K inhibition. (A) In vitro drug sensitivity assays with the PI3K inhibitor LY204002 were set up using synchronous ring stage parasites and % growth was measured 72 h later. Shown are the average IC₅₀ values from at least 3 independent biological replicates. A two tailed paired t test was performed for statistical comparison. (B, i) Western blot from a representative experiment showing the effect of LY294002 treatment on parasite-associated Hb levels. Trophozoite (3D7) and ring/trophozoite (PfPX1-KOs) stages were treated with 100 μM LY294002 for 4 h. HSP70 was used as a loading control for normalization. Results from one experiment representative of 3 independent biological replicates shown. (B, ii) Quantification of the increase in parasite-associated Hb normalized to HSP70 upon LY294002 treatment from Western blot by densitometry analyses of three independent biological replicates. Unpaired t tests were performed for statistical evaluation. (C, i) Quantification of the increase in DV proximal and total Hb-containing vesicles upon LY294002 treatment. IFA using an anti-Hb antibody was performed on parasite lines at trophozoite (3D7) and ring/trophozoite (PfPX1-KO-1) stages treated with 100 μM LY294002 for 4 h. Mean ± SD from 2 independent biological replicates is shown. (C, ii) Representative image of a DV distal vesicle. (C, iii) Representative image of a DV proximal vesicle. White arrows: Highlighting the Hb vesicles. Dapi: parasite nuclei.