Plug for the parasitophorous duct: a solution of two conundra

In order to solve this apparent conundrum, we propose a parasitophorous duct that is sealed by a “plug” composed of a protein complex, impermeable to all solutes but containing transporters for entry of a select set of serum proteins, i.e. a modified tight junction. This plug originates from the mechanism of merozoite ingress into RBC, a process initiated by formation of an attachment complex between the apical end of an invading merozoite and host cell surface, first reported through painstaking analysis of transmission electron micrographs by Aikawa et al. [6], followed by movement of this attachment complex (in the form of a belt-like tight junction) over the merozoite surface towards the posterior end, driven by an actin-myosin motor system located beneath the merozoite plasma membrane and connected to the tight junction endodomains [7]. When the tight junction complex converges at the merozoite posterior region, it is believed that the tight junction complex disassembles allowing the apposed RBC plasma membranes to fuse and generate two separate intact membranes, namely, PVM and RBCM. Applying Occam’s razor, we posit that the tight junction positioned at the RBC surface remains intact, interacting with the two apposed RBCMs, thus forming the plug, but allows detachment of the merozoite to reside and develop within the PV (Fig. 1). The tight junction transmembrane domains would prevent any lateral diffusion of integral membrane proteins across this barrier, consistent of the presence of distinct sets of PVM and RBCM proteins [8, 9]. This model allows normal lateral movement of lipids as expected of a continuous bilayer. Once an external protein gains entry to the PV, its distribution within the tubovesicular network (TVN) and Maurer’s clefts (formed by budding from TVN [10]) is assured [2, 5]. If a protein transporter exists in the parasitophorous tight junction pore structure, might not a similar transporter exist on the merozoite plasma membrane, thereby affording an explanation for the appearance of serum protein(s) in parasite cytosol [1, 2, 5]? As regards the selectivity and specificity of the putative protein transporter(s), it is worth noting the internalized serum proteins vitamin K-dependent protein S and prothrombin have a γ-carboxyglutamic acid-rich domain and vitronectin that does not contain this domain forms a complex with thrombin in serum [11], and plasminogen is found on infected RBC surface and could be a bystander in the uptake process as its presence in internalized parasite is 40- and 100-folds less than that of prothrombin and vitamin K-dependent protein S respectively [2].

The presence of an intact discrete PVM poses the second conundrum: requirement of PVM lysis followed by that of infected RBCM for merozoite egress, an event requiring activation of a cascade of parasite proteases [12], but the exact mechanism of dissolution of the two lipid bilayers by protease(s) remains unclear. This conundrum is readily solved by the existence of the protein plug, degradation/destabilization of which causes restoration of the pore opening that enlarges through lateral sheer stress exerted by the elastic infected RBCM [13], thereby raising the PVM to RBC surface and expelling the progeny merozoites (probably also aided by osmotic pressure) through an apparent opening and in a fixed trajectory. This phenomenon of merozoites’ egress has been directly visualized using high-speed differential interference contrast video microscopy and epifluorescence [14]. Subsequent shearing of the PVM fabric would account for the infected RBCM contortions observed at the edge of the egress opening [15]. The model does not exclude the distinct possibility that the porin-containing PVM [16] is ruptured (by osmotic pressure) during but not before merozoites’ egress through the opening in RBCM, and given the rapidity of the egress events (a matter of seconds [16, 17]) it may appear that PVM rupture occurs prior to merozoite expulsion.

Is there any evidence for the existence of this conjectural parasitophorous duct plug? Baum et al. [18] have recounted the steps leading to obtaining a high-resolution 3-dimensional image of a merozoite invading RBC, which depicts the progression starting with a small apical tight junction that then moves to encircle the merozoite and finally seals the entry at the posterior end. It remains to be shown if this tight junction plug remains at infected RBCM blocking the entrance of the parasitophorous pore. We believe this is a testable model though application of such imaging tools as cryo-X-ray tomography and super-resolution fluorescence microscopy, which already have been applied to the invasion process [19].