Selective Inhibition of a Two-step Egress of Malaria Parasites from the Host Erythrocyte

Mark E Wickham,Janetta G Culvenor,Alan F Cowman

doi:10.1074/jbc.m305252200

Abstract

Escape from the host erythrocyte by the invasive stage of the malaria parasite Plasmodium falciparum is a fundamental step in the pathogenesis of malaria of which little is known. Upon merozoite invasion of the host cell, the parasite becomes enclosed within a parasitophorous vacuole, the compartment in which the parasite undergoes growth followed by asexual division to produce 16-32 daughter merozoites. These daughter cells are released upon parasitophorous vacuole and erythrocyte membrane rupture. To examine the process of merozoite release, we used P. falciparum lines expressing green fluorescent protein-chimeric proteins targeted to the compartments from which merozoites must exit: the parasitophorous vacuole and the host erythrocyte cytosol. This allowed visualization of merozoite release in live parasites. Herein we provide the first evidence in live, untreated cells that merozoite release involves a primary rupture of the parasitophorous vacuole membrane followed by a secondary rupture of the erythrocyte plasma membrane. We have confirmed, with the use of immunoelectron microscopy, that parasitophorous vacuole membrane rupture occurs before erythrocyte plasma membrane rupture in untransfected wild-type parasites. We have also demonstrated selective inhibition of each step in this two-step process of exit using different protease inhibitors, implicating the involvement of distinct proteases in each of these steps. This will facilitate the identification of the parasite and host molecules involved in merozoite release.

Highlights

During the erythrocytic phase of the Plasmodium falciparum life cycle, the merozoites released initially from infected hepatocytes attach to and invade human erythrocytes in the bloodstream
Ultrastructural evidence suggests that during schizogony, the parasite plasma membrane invaginates to surround the merozoites forming within the confines of the parasitophorous vacuole [17] and that late in schizogony, the parasitophorous vacuole membrane may be absent, with the fully formed merozoites free within the host erythrocyte [17, 18]
Intraerythrocytic Parasitophorous Vacuole Rupture in GFPexpressing Parasites—To investigate the sequence of events involved in schizont rupture, transgenic P. falciparum lines have been generated that traffic GFP fusions to the compartments that the parasite must traverse upon exit: the parasitophorous vacuole and the host erythrocyte cytosol

Summary

Introduction

During the erythrocytic phase of the Plasmodium falciparum life cycle, the merozoites released initially from infected hepatocytes attach to and invade human erythrocytes in the bloodstream. Intraerythrocytic Parasitophorous Vacuole Rupture in GFPexpressing Parasites—To investigate the sequence of events involved in schizont rupture, transgenic P. falciparum lines have been generated that traffic GFP fusions to the compartments that the parasite must traverse upon exit: the parasitophorous vacuole and the host erythrocyte cytosol.

Results

Conclusion