Plasmodium falciparum AMA1 binds a rhoptry neck protein homologous to TgRON4, a component of the moving junction in Toxoplasma gondii

Abstract

Plasmodium falciparum apical membrane antigen 1 (PfAMA1) coimmunoprecipitates with the Plasmodium homologue of TgRON4, a secreted rhoptry neck protein of Toxoplasma gondii that migrates at the moving junction in association with TgAMA1 during invasion. PfRON4 also originates in the rhoptry necks, suggesting that this unusual collaboration of micronemes and rhoptries is a conserved feature of Apicomplexa.

FIG. 1.

Identification of PfAMA1-associated proteins. A. MAb 28G2-affinity-selected proteins from RIPA detergent extracts of segmented schizonts were separated on reducing SDS-PAGE gels and stained with Coomassie blue. Molecular mass markers (kDa) are denoted on the left, and the sizes of the major bands are indicated on the right (arrows) and are based on relative mobility. The profiles for the RIPA parasite extract (7 × 10⁶ parasite equivalents) before (lane 1) and after (lane 2) immuno-selection are shown. MAb 28G2-immuno-selected proteins (7 × 10⁸ parasite equivalents) are shown (lane 3) in comparison with those precipitated by protein G-Sepharose beads alone (lane 4). B. Immunoblot of parasite extract and affinity-selected proteins probed with anti-PfAMA1 MAb 28G2. Note that this is a different gel from that shown in panel A, in order to better resolve proteins in the 50- to 100-kDa range. C. Immunoblot of the lanes shown in panel B that were probed with anti-PfRON4 MAb 24C6.

FIG. 2.

PfRON4 localizes to the rhoptry necks. Immuno-electron microscopy with MAb 24C6 and protein A-gold shows specific labeling in the rhoptry necks of P. falciparum schizonts. Infected human erythrocytes were embedded in LRWhite and stained with MAb 24C6 as previously described (26). The enlarged area shows a longitudinally sectioned rhoptry with PfRON4 labeling restricted to the narrow apical end of the rhoptries in a maturing merozoite (arrows).