Multiprotein complex between the GPI-anchored CyRPA with PfRH5 and PfRipr is crucial for Plasmodium falciparum erythrocyte invasion

Abstract

Erythrocyte invasion by Plasmodium falciparum merozoites is a highly intricate process in which Plasmodium falciparum reticulocyte binding-like homologous protein 5 (PfRH5) is an indispensable parasite ligand that binds with its erythrocyte receptor, Basigin. PfRH5 is a leading blood-stage vaccine candidate because it exhibits limited polymorphisms and elicits potent strain-transcending parasite neutralizing antibodies. However, the mechanism by which it is anchored to the merozoite surface remains unknown because both PfRH5 and the PfRH5-interacting protein (PfRipr) lack transmembrane domains and GPI anchors. Here we have identified a conserved GPI-linked parasite protein, Cysteine-rich protective antigen (CyRPA) as an interacting partner of PfRH5-PfRipr that tethers the PfRH5/ PfRipr/CyRPA multiprotein complex on the merozoite surface. CyRPA was demonstrated to be GPI-linked, localized in the micronemes, and essential for erythrocyte invasion. Specific antibodies against the three proteins successfully detected the intact complex in the parasite and coimmunoprecipitated the three interacting partners. Importantly, full-length CyRPA antibodies displayed potent straintranscending invasion inhibition, as observed for PfRH5. CyRPA does not bind with erythrocytes, suggesting that its parasite neutralizing antibodies likely block its critical interaction with PfRH5-PfRipr, leading to a blockade of erythrocyte invasion. Further, CyRPA and PfRH5 antibody combinations produced synergistic invasion inhibition, suggesting that simultaneous blockade of the PfRH5-Basigin and PfRH5/PfRipr/CyRPA interactions produced an enhanced inhibitory effect. Our discovery of the critical interactions between PfRH5, PfRipr, and the GPI-anchored CyRPA clearly defines the components of the essential PfRH5 adhesion complex for P. falciparum erythrocyte invasion and offers it as a previously unidentified potent target for antimalarial strategies that could abrogate formation of the crucial multiprotein complex. malaria | erythrocyte invasion | protein-protein interactions | blood-stage vaccines | PfRH5

Evaluating antidisease immunity to malaria and implications for vaccine design

“…Furthermore, even at such high antibody levels, growth inhibition is rarely absolute. Antibodies against cysteine‐rich protective antigen (CyRPA) (full length) in two‐cycle invasion assays showed 85% inhibition at 10 mg/ml,69 antibodies against Rh5 (full‐length) showed 83% inhibition in a one‐cycle invasion assay at 10 mg/ml 42,70 and a 40% invasion inhibition was observed for antibodies against Rh5 interacting protein (Ripr) (ectodomain) at 2 mg/ml in a one‐cycle assay 42. Similarly, antibodies against several other characterized antigens (such as MSPDBL‐1 and ‐2)71, 72 showed a comparable trend of invasion inhibition and require fairly large antibody concentrations.…”

Structurally conserved erythrocyte-binding domain in Plasmodium provides a versatile scaffold for alternate receptor engagement

“…This receptor-adhesin pair mediates an essential molecular event during parasite entry, and sequence polymorphisms within PfRh5 and/or basigin have been shown to modulate host tropism (16,17). The addition of antibodies against PfRh5, basigin, or PfRh5-interacting partners (PfRipr and PfCyRPA) in P. falciparum growth assays results in the strong inhibition of parasite growth across multiple strains (15,(18)(19)(20)(21). Further promising results show that Aotus nancymaae monkeys immunized with the anti-PfRh5 vaccine are protected against severe infection (22).…”

Human vaccination against RH5 induces neutralizing antimalarial antibodies that inhibit RH5 invasion complex interactions

“…Nevertheless, substantial progress has been made in recent years, with the identification of a new generation of merozoite antigen targets that are essential and that exhibit relatively low levels of polymorphism, leading to induction of strain-transcending antibodies by vaccination in preclinical models (11). Three of these targets form a complex, including the P. falciparum reticulocyte-binding protein homolog 5 (RH5) (12), the cysteine-rich protective antigen (CyRPA) (13), and the RH5-interacting protein (Ripr) (14). Of these, vaccine development efforts are currently most advanced for RH5 (15).…”