Loss of PfGAP40 affects targeting of PfGAP45 and PfGAP50 to the cytoskeletal IMC. (A) IP analysis of PfGAP40cKD parasites. The PfGAP40cKD parasites expressing PfGAP40-Ty were subjected to IP with the anti-Ty antibody, and the presence of PfGAP40-Ty (left), PfGAP45 (middle), and PfGAP50 (right) in the elution was checked by Western blotting. T, total protein; S, supernatant of co-IP analysis; P, pellet of co-IP analysis. Arrows indicate the relative endogenous protein. (B) Representative IFA images of PfGAP45 and PfGAP50 in mock- (No-GlcN) or GlcN-treated PfGAP40cKD parasites. BF, bright field; blue, nucleus; green, rabbit anti-PfGAP45 or anti-PfGAP50. Scale bars: 5 μm. (C) Western blot analysis of PfGAP40 KD parasites shows the dominant expression of glideosome-associated proteins PfGAP40, PfGAP45, and PfGAP50 in matured schizonts (left) and a dramatic reduction of PfGAP45 and PfGAP50 expression in schizonts upon PfGAP40 knockdown (right). PfGAP40cKD parasites were tightly synchronized and harvested at 0, 6, 12, 24, 36, 40, and 44 hpi, following mock or GlcN treatment and incubated with anti-PfGAP45, anti-PfGAP50, anti-Ty, and anti-aldolase antibodies. Molecular mass in kDa. Representative results from three biological replicates are shown. He L, Qiu Y, Pang G, Li S, Wang J, Feng Y, Chen L, Zhu L, Liu Y, Cui L, Cao Y, Zhu X. Plasmodium falciparum GAP40 Plays an Essential Role in Merozoite Invasion and Gametocytogenesis. Microbiol Spectr. 2023 11(3):e0143423.. PMID: 37249423