Proteomic Analysis of the Plasmodium berghei Gametocyte Egressome and Vesicular bioID of Osmiophilic Body Proteins Identifies Merozoite TRAP-like Protein (MTRAP) as an Essential Factor for Parasite Transmission

Abstract

Malaria transmission from an infected host to the mosquito vector requires the uptake of intraerythrocytic sexual precursor cells into the mosquito midgut. For the release of mature extracellular gametes two membrane barriers-the parasite parasitophorous vacuole membrane and the host red blood cell membrane-need to be dissolved. Membrane lysis occurs after the release of proteins from specialized secretory vesicles including osmiophilic bodies. In this study we conducted proteomic analyses of the P. berghei gametocyte egressome and developed a vesicular bioID approach to identify hitherto unknown proteins with a potential function in gametocyte egress. This first Plasmodium gametocyte egressome includes the proteins released by the parasite during the lysis of the parasitophorous vacuole membrane and red blood cell membrane. BioID of the osmiophilic body protein MDV1/PEG3 revealed a vesicular proteome of these gametocyte-specific secretory vesicles. Fluorescent protein tagging and gene deletion approaches were employed to validate and identify a set of novel factors essential for this lysis and egress process. Our study provides the first in vivo bioID for a rodent malaria parasite and together with the first Plasmodium gametocyte egressome identifies MTRAP as a novel factor essential for mosquito transmission. Our data provide an important resource for proteins potentially involved in a key step of gametogenesis. Molecular & Cellular

Plasmodium Merozoite TRAP Family Protein Is Essential for Vacuole Membrane Disruption and Gamete Egress from Erythrocytes

“…Activated male Pb MTRAP KO gametocytes formed motile flagella, but they remained intracellular, beating as a thick bundle of flagella, suggesting that they were unable to egress from the PV or the host erythrocyte (Figure 3B; Movie S1). A similar conclusion of lack of egress of P. berghei MTRAP KO was reported in a recently published paper (Kehrer et al., 2016). …”

Motility precedes egress of malaria parasites from oocysts

“…Proteins containing TSRs are widespread among animals and protozoans and are mostly extracellular or secreted proteins (Tucker, 2004). Also, at least one TSR is contained within all proteins of the TRAP-family, including MTRAP, which was recently found to be important for gametocyte egress from red blood cells (Kehrer et al, 2016a; Bargieri et al, 2016; Morahan et al, 2009) (Figure 1A). In a search for uncharacterized TRAP-family-like TSR-containing proteins, we found a protein with unknown function in the rodent model parasite P. berghei (PBANKA_0707900).…”

“…However, the number of non-related proteins that function in egress, and the lack of any clear interaction between these proteins, suggests that even more proteins are involved in the egress of sporozoites from oocysts. It was recently shown that the TRAP-family member MTRAP, previously thought to be important for red blood cell invasion by merozoites, is crucial for the egress of gametocytes from host cells (Kehrer et al, 2016a; Bargieri et al, 2016). We rationalized that similar proteins might also play a role in oocyst egress and searched for distantly related TRAP-like proteins.…”

BioID as a Tool for Protein-Proximity Labeling in Living Cells

“…Since its development and initial application in 2012 (7), BioID has been cited and/or applied in over 200 PubMed-published articles and has been utilized in several unicellular organisms (see (12–15) for examples), mammalian cells (5), plants (16,17), and mice (18–20), including compartmental proteomics of a parasitic organism infecting mice (21). Novel or improved applications using BioID ligase have spurred numerous follow-up articles including a smaller version of BioID with improved sensitivity and localization (22), its use for identifying protein-RNA interactions (23), split-BioID studies (24,25), and faster versions of BioID (26).…”