SummaryApical membrane antigen-1 (AMA1) is a conserved apicomplexan protein that plays an important but undefined role in host cell invasion. We have studied the fate of Plasmodium falciparum AMA1 (PfAMA1) during erythrocyte invasion by the malaria merozoite, and compared it with that of the Toxoplasma gondii orthologue, TgAMA1. Shedding of the PfAMA1 ectodomain goes essentially to completion during invasion, and occurs predominantly or exclusively via juxtamembrane cleavage at the previously identified sheddase cleavage site, Thr517. Only the resulting juxtamembrane stub of the ectodomain is efficiently carried into the host cell, and this remains distributed around the plasma membrane of the intracellular ringstage parasite. Inhibition of normal shedding, however, results in proteolysis at an intramembrane, rhomboid-like cleavage site, and PfAMA1 is susceptible to cleavage by Drosophila rhomboid-1, showing that it can be a substrate for intramembrane cleavage but is not normally processed in this manner. In contrast, shedding of TgAMA1 from the surface of extracellular tachyzoites occurs exclusively via cleavage within the luminal half of its transmembrane domain by a rhomboid-like protease. Also unlike PfAMA1, complete TgAMA1 shedding does not accompany Toxoplasma invasion as the intact protein was readily detected on the surface of newly invaded tachyzoites. This work reveals unexpected differences in the manner in which Plasmodium and Toxoplasma shed AMA1 from the surface of invasive zoites, and demonstrates the presence at the malaria merozoite surface of a rhomboid-like protease.

Many intracellular pathogens are separated from the cytosol of their host cells by a vacuole membrane. This membrane serves as a critical interface between the pathogen and the host cell, across which nutrients are imported, wastes are excreted, and communication between the two cells takes place. Very little is known about the vacuole membrane proteins mediating these processes in any host-pathogen interaction. During a screen for monoclonal antibodies against novel surface or secreted proteins of Toxoplasma gondii, we identified ROP4, a previously uncharacterized member of the ROP2 family of proteins. We report here on the sequence, posttranslational processing, and subcellular localization of ROP4, a type I transmembrane protein. Mature, processed ROP4 is localized to the rhoptries, secretory organelles at the apical end of the parasite, and is secreted from the parasite during host cell invasion. Released ROP4 associates with the vacuole membrane and becomes phosphorylated in the infected cell. Similar results are seen with ROP2. Further analysis of ROP4 showed it to be phosphorylated on multiple sites, a subset of which result from the action of either host cell protein kinase(s) or parasite kinase(s) activated by host cell factors. The localization and posttranslational modification of ROP4 and other members of the ROP2 family of proteins within the infected cell make them well situated to play important roles in vacuole membrane function.Toxoplasma gondii is an obligate intracellular parasite capable of invading nearly any nucleated cell within its vertebrate hosts. The process of host cell invasion is similar in all apicomplexan parasites, including T. gondii, Plasmodium spp. (causative agents of malaria), Eimeria tenella, and Cryptosporidium parvum. Internalized parasites undergo several rounds of replication within the parasitophorous vacuole (PV) and ultimately lyse the host cell. Repeated cycles of host cell invasion and lysis are directly responsible for the disease caused by T. gondii, which is particularly severe in immunocompromised individuals and in the congenitally infected fetus (19,23,33).As with all apicomplexan parasites, a central feature of host cell invasion by T. gondii tachyzoites is the sequential release of proteins from apical secretory organelles. The tachyzoite secretes proteins first from the micronemes, then from the rhoptries, as it pulls itself into the host cell using a myosin-based motor complex (9,12,24). Proteins released from the micronemes include a variety of adhesins, which are thought to directly mediate binding to ligands on the host cell surface (reviewed in references 43 and 46). Rhoptry proteins are released concurrent with the formation of the PV and are thought to contribute to both the formation and functional properties of the PV membrane (PVM) (3, 9, 12, 41).Host cell invasion by T. gondii tachyzoites is inhibited by cytochalasin D, while host cell attachment and rhoptry secretion remain unaffected (9,14). Under these conditions, the contents of the rhoptrie...

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Chronic granulomatous disease (CGD), a primary immunodeficiency characterized by a deficient neutrophil oxidative burst and the inadequate killing of microbes, is well known to cause a significantly increased risk of invasive infection. However, infectious complications are not the sole manifestations of CGD; substantial additional morbidity is driven by noninfectious complications also. These complications can include, for example, a wide range of inflammatory diseases that affect the gastrointestinal tract, lung, skin, and genitourinary tract and overt autoimmune disease. These diseases can occur at any age and are especially problematic in adolescents and adults with CGD. Many of these noninfectious complications present a highly challenging therapeutic conundrum, wherein immunosuppression must be balanced against an already markedly increased risk of invasive fungal and bacterial infections. In this review, the myriad noninfectious complications of CGD are discussed, as are important gaps in our understanding of these processes, which warrant further investigation.