Super-Resolution Dissection of Coordinated Events during Malaria Parasite Invasion of the Human Erythrocyte

Abstract

Erythrocyte invasion by the merozoite is an obligatory stage in Plasmodium parasite infection and essential to malaria disease progression. Attempts to study this process have been hindered by the poor invasion synchrony of merozoites from the only in vitro culture-adapted human malaria parasite, Plasmodium falciparum. Using fluorescence, three-dimensional structured illumination, and immunoelectron microscopy of filtered merozoites, we analyze cellular and molecular events underlying each discrete step of invasion. Monitoring the dynamics of these events revealed that commitment to the process is mediated through merozoite attachment to the erythrocyte, triggering all subsequent invasion events, which then proceed without obvious checkpoints. Instead, coordination of the invasion process involves formation of the merozoite-erythrocyte tight junction, which acts as a nexus for rhoptry secretion, surface-protein shedding, and actomyosin motor activation. The ability to break down each molecular step allows us to propose a comprehensive model for the molecular basis of parasite invasion.

Minimal requirements for actin filament disassembly revealed by structural analysis of malaria parasite actin-depolymerizing factor 1

“…Free merozoites were isolated and processed for microscopy following ref. 24. Antibodies and image processing are described in SI Materials and Methods.…”

“…Wild-type (D10) asexual parasites were maintained in standard culture conditions and synchronized as described (24). Free merozoites were isolated and processed for microscopy following ref.…”

“…We explored PfADF1 localization by IFA in free merozoites captured mid-invasion and colabeled with rhoptry neck protein 4 (RON4), a marker of the tight junction (24) that is the point of close apposition between host and parasite membranes during invasion. The tight junction was shown recently to be the site at which a ring of actin forms (24), a feature consistent with engagement of the actomyosin motor at the junction. PfADF1 retained its cytosolic localization during invasion ( Fig.…”

Regulation of Plasmodium falciparum Development by Calcium-dependent Protein Kinase 7 (PfCDPK7)

“…The growth and morphology of parasites post-invasion was monitored by IFA for RAP1/2 and MSP1, which are located in PV and PVM, respectively (23). The rhoptry bulb proteins such as the low molecular weight rhoptry bulb complex proteins RAP1 and 2 are transferred to the PV postinvasion (23), (24).…”

Un complexe moléculaire unique à l’origine de la motilité et de l’invasion des Apicomplexes

“…Une fois sécrétée, AMA1 reste insérée dans la membrane plasmique du parasite et migre au niveau de la jonction mobile durant l'invasion. Cette protéine apparaît comme un anneau à l'intérieur du cercle formé par RON4 lors de l'invasion de mérozoïtes de P. falciparum [38]. Les trois protéines RON sont, elles, localisées au-delà de la membrane plasmique de la cellule hôte où elles pourraient interagir avec le cytosquelette cellulaire [39].…”