Mononeme: A new secretory organelle in Plasmodium falciparum merozoites identified by localization of rhomboid-1 protease

Abstract

Compartmentalization of proteins into subcellular organelles in eukaryotic cells is a fundamental mechanism of regulating complex cellular functions. Many proteins of Plasmodium falciparum merozoites involved in invasion are compartmentalized into apical organelles. We have identified a new merozoite organelle that contains P. falciparum rhomboid-1 (PfROM1), a protease that cleaves the transmembrane regions of proteins involved in invasion. By immunoconfocal microscopy, PfROM1 was localized to a single, thread-like structure on one side of the merozoites that appears to be in close proximity to the subpellicular microtubules. PfROM1 was not found associated with micronemes, rhoptries, or dense granules, the three identified secretory organelles of invasion. Release of merozoites from schizonts resulted in the movement of PfROM1 from the lateral asymmetric localization to the merozoite apical pole and the posterior pole. We have named this single thread-like organelle in merozoites, the mononeme. malaria A picomplexa are named for a set of secretory organelles (rhoptries, micronemes, and dense granules) found at the apical end of these parasites, the end that invades cells in the vertebrate host (1). These organelles were first identified by their distinct morphological appearances in transmission electron microscopy (2-4). Many parasite proteins required for invasion of erythrocytes are segregated into the micronemes (5, 6). For example, Plasmodium falciparum apical membrane antigen 1 (AMA1) is held in the micronemes in merozoites inside of erythrocytes. Release from the erythrocyte triggers the movement of AMA1 to the cell surface where it functions in invasion (7,8). Similarly, rhoptries sequester a different set of proteins and their contents are released onto the erythrocyte surface (4, 9), presumably to break the local cytoskeleton and to initiate formation of the parasitophorous vacuole. Taken together, these observations provide evidence for compartmentalization of parasite proteins into distinct organelles with related functions in invasion. Presumably, such compartmentalization provides a mechanism for orchestrating the timing of delivery or activity of the proteins during the complex process of invasion.Apicomplexan rhomboid proteases have been implicated to play an important role in host cell invasion (10, 11). PfROM1 substrates have been identified in P. falciparum by using a mammalian cell-based proteolytic assay (12) that identified various micronemal proteins as potential substrates including AMA1. Because AMA1 has been implicated as essential for invasion, separation of PfROM1 from this substrate within the parasite may be important to prevent premature cleavage. Studies defining the spatiotemporal distribution of PfROM1 in P. falciparum merozoites are, therefore, likely to contribute to a clearer understanding of its role in erythrocyte invasion. Expression of hemagglutinin (HA)-tagged P. falciparum rhomboid-1 (PfROM1) localizes it to a new subcellular compartment distinct from other known meroz...

Analysis of the Conformation and Function of the Plasmodium falciparum Merozoite Proteins MTRAP and PTRAMP

“…Immunofluorescence assay (IFA) analysis was performed as per the standard protocol (26,33). In brief, schizont-enriched parasites were smeared on slides and stored at Ϫ80°C.…”

New insights into parasite rhomboid proteases

“…TgROM1 is a micronemal protein, which is catalytically active in vitro against Spitz, but its physiological substrate(s) are unknown [52,53]. In P. falciparum, ROM1 has been localized to either a new secretory organelle called mononeme [54] or to the micronemes [48]. PfROM1 is an active protease but, again, its physiological substrates remain to be identified [50].…”

“…PbROM1PfROM1PyROM1 Micronemes (PyROM1/PfROM1/PbROM1) [48,56,57] Mononemes (PfROM1) [54] In vitro DmSpitz/PfAMA1/PfRH1/PfRh2B/PfMAEBL/PyUIS4 [50,57] Parasitophorous vacuole modification (PyROM1) Parasite development (PbROM1) Unknown (PfROM1) In vivo: unknown PfROM4…”

Ubiquitylation-Dependent Negative Regulation of WASp Is Essential for Actin Cytoskeleton Dynamics

“…Pearson's colocalization correlation coefficient analysis was performed as previously described (33). Imaris 7.0 software (Bitplane AG, Zurich, Switzerland) was used to distinguish between actual colocalization and random association between signals from two fluorescent labels over the entire three-dimensional (3D) image.…”