Truncation of RhopH3 leads to mistrafficking of components of the RhopH complex and loss of complex formation. A) IFA showing colocalization of RhopH3, RhopH2 and RhopH1/Clag3.1 with the rhoptry marker RAP2 in schizonts of control (DMSO) rhopH3-loxP parasites but loss of colocalization following rapamycin (Rapa) treatment. Parasite nuclei were visualized by staining with 4,6-diamidino-2-phenylindole (DAPI). Scale bar, 5 μm. Immunofluorescence analysis (IFA) showed that, as expected, RhopH3 colocalized with the rhoptry marker RAP2 in mature schizonts of control rhopH3-loxP parasites. However, in rapamycin-treated (RhopH3Δ4-219 6) parasites, this colocalization was lost, although RAP2 was still detected in a punctate, apically-disposed pattern typical of rhoptries B) Colocalization of the members of the RhopH complex. RhopH3, RhopH2 and Rhop 1/Clag3.1 colocalize in rhopH3-loxP parasites treated with DMSO, but this colocalization is lost in parasites treated with rapamycin. Scale bar, 2 μm. Neither RhopH2 nor RhopH1/Clag3.1 colocalized with 233 RhopH3Δ4-6 in the mutant parasites. The RhopH2 and RhopH1/Clag3.1 signals were also distinct in the mutant parasites, although in this case some limited colocalization of these proteins was apparent (bottom images).

Sherling ES, Knuepfer E, Brzostowski JA, Miller LH, Blackman MJ, van Ooij C. The Plasmodium falciparum rhoptry protein RhopH3 plays essential roles in host cell invasion and nutrient uptake. Elife. 2017 6. pii: e23239.