Dissecting the interface between apicomplexan parasite and host cell: Insights from a divergent AMA–RON2 pair

Abstract

Plasmodium falciparum and Toxoplasma gondii are widely studied parasites in phylum Apicomplexa and the etiological agents of severe human malaria and toxoplasmosis, respectively. These intracellular pathogens have evolved a sophisticated invasion strategy that relies on delivery of proteins into the host cell, where parasite-derived rhoptry neck protein 2 (RON2) family members localize to the host outer membrane and serve as ligands for apical membrane antigen (AMA) family surface proteins displayed on the parasite. Recently, we showed that T. gondii harbors a novel AMA designated as TgAMA4 that shows extreme sequence divergence from all characterized AMA family members. Here we show that sporozoite-expressed TgAMA4 clusters in a distinct phylogenetic clade with Plasmodium merozoite apical erythrocyte-binding ligand (MAEBL) proteins and forms a high-affinity, functional complex with its coevolved partner, TgRON2 L1 . High-resolution crystal structures of TgAMA4 in the apo and TgRON2 L1 -bound forms complemented with alanine scanning mutagenesis data reveal an unexpected architecture and assembly mechanism relative to previously characterized AMA-RON2 complexes. Principally, TgAMA4 lacks both a deep surface groove and a key surface loop that have been established to govern RON2 ligand binding selectivity in other AMAs. Our study reveals a previously underappreciated level of molecular diversity at the parasite-host-cell interface and offers intriguing insight into the adaptation strategies underlying sporozoite invasion. Moreover, our data offer the potential for improved design of neutralizing therapeutics targeting a broad range of AMA-RON2 pairs and apicomplexan invasive stages.Apicomplexa | Toxoplasma gondii | invasion | moving junction | X-ray crystallography P hylum Apicomplexa comprises >5,000 parasitic protozoan species, many of which cause devastating diseases on a global scale. Two of the most prevalent species are Toxoplasma gondii and Plasmodium falciparum, the causative agents of toxoplasmosis and severe human malaria, respectively (1, 2). The obligate intracellular apicomplexan parasites lead complex and diverse lifestyles that require invasion of many different cell types. Despite this diversity of target host cells, most apicomplexans maintain a generally conserved mechanism for active invasion (3). The parasite initially glides over the surface of a host cell and then reorients to place its apical end in close contact to the hostcell membrane. After this initial attachment, a circumferential ring of adhesion (termed the moving or tight junction) is formed, through which the parasite actively propels itself while concurrently depressing the host-cell membrane to create a nascent protective vacuole (4).Formation of the moving junction relies on a pair of highly conserved parasite proteins: rhoptry neck protein 2 (RON2) and apical membrane antigen 1 (AMA1). Initially, parasites discharge RON2 into the host cell membrane where an extracellular portion (domain 3; D3) serves as a ligand for AMA1 displa...

AMA1 and MAEBL are important forPlasmodium falciparumsporozoite infection of the liver

“…is the chimeric protein MAEBL (Kappe et al, 1998). While AMA1 and MAEBL are not identical, they share similarity and are evolutionarily related (Kappe et al, 1998;Parker et al, 2016). MAEBL-deficient P. berghei and P. falciparum sporozoites show a striking defect during invasion of mosquito salivary glands (Kariu et al, 2002;Saenz et al, 2008), a finding that was also confirmed in our study.…”

“…MAEBL is conserved in Plasmodium spp. and clusters phylogenetically with AMA4 of Toxoplasma gondii (Parker et al, 2016). While MAEBL can bind erythrocytes (Kappe et al, 1998), genetic disruption had no effect on blood-stage growth, indicating it is not solely essential for erythrocyte invasion (Fu et al, 2005;Kariu, Yuda, Yano, & Chinzei, 2002;Saenz, Balu, Smith, Mendonca, & Adams, 2008).…”

Genetic Analysis of Babesia Isolates from Cattle with Clinical Babesiosis in Sri Lanka

“…The 371-bp ama-1 sequences from cows R and V (GenBank accession number LC385894) were identical to each other and to the long ama-1 gene fragment initially isolated from cow R (GenBank accession number LC385893). Phylogenetic trees based on the ama-1 gene were used to investigate the evolutionary relationships of apicomplexan parasites, such as species of Plasmodium and Babesia (61,(63)(64)(65). In a phylogenetic construction used to investigate the positions of Babesia sp.…”

Gliding motility powers invasion and egress in Apicomplexa

“…Concordantly, genetic approaches in T. gondii have validated the contribution of the AMA1-RONs complex to invasion [133][134][135][136] , although parasite motility was not substantially affected (Supplementary information S6 (table)). Although a TgAMA1-null mutant could be obtained in vitro 137 , the upregulation of TgAMA1 homologues that sustain residual invasion confirmed the crucial role of the moving junction in host cell penetration 134,138 . In Plasmodium spp., it was not possible to knockout AMA1 and RON4; however, the conditional depletion of P. berghei AMA1 (PbAMA1) and PfAMA1 caused a defect in merozoite invasion 139,140 and in the sealing of the parasitophorous vacuole at the end of invasion 140 .…”