Temporal Expression and Localization Patterns of Variant Surface Antigens in Clinical Plasmodium falciparum Isolates during Erythrocyte Schizogony

Abstract

Avoidance of antibody-mediated immune recognition allows parasites to establish chronic infections and enhances opportunities for transmission. The human malaria parasite Plasmodium falciparum possesses a number of multi-copy gene families, including var, rif, stevor and pfmc-2tm, which encode variant antigens believed to be expressed on the surfaces of infected erythrocytes. However, most studies of these antigens are based on in vitro analyses of culture-adapted isolates, most commonly the laboratory strain 3D7, and thus may not be representative of the unique challenges encountered by P. falciparum in the human host. To investigate the expression of the var, rif-A, rif-B, stevor and pfmc-2tm family genes under conditions that mimic more closely the natural course of infection, ex vivo clinical P. falciparum isolates were analyzed using a novel quantitative real-time PCR approach. Expression patterns in the clinical isolates at various time points during the first intraerythrocytic developmental cycle in vitro were compared to those of strain 3D7. In the clinical isolates, in contrast to strain 3D7, there was a peak of expression of the multi-copy gene families rif-A, stevor and pfmc-2tm at the young ring stage, in addition to the already known expression peak in trophozoites. Furthermore, most of the variant surface antigen families were overexpressed in the clinical isolates relative to 3D7, with the exception of the pfmc-2tm family, expression of which was higher in 3D7 parasites. Immunofluorescence analyses performed in parallel revealed two stage-dependent localization patterns of RIFIN, STEVOR and PfMC-2TM. Proteins were exported into the infected erythrocyte at the young trophozoite stage, whereas they remained inside the parasite membrane during schizont stage and were subsequently observed in different compartments in the merozoite. These results reveal a complex pattern of expression of P. falciparum multi-copy gene families during clinical progression and are suggestive of diverse functional roles of the respective proteins.

Plasmodium falciparum STEVOR phosphorylation regulates host erythrocyte deformability enabling malaria parasite transmission

“…We further confirmed these observations in 22-hpi asexual stages before endogenous expression of STEVOR. 16,30,31 At these stages, overexpression of the full copy of STEVOR in the Full-Ty1 line substantially increased the retention rates compared with the control line (78% vs 51.9%, P 5 0), whereas the retention rates of the Trunc-Ty1 line were at the same level as the control line (54.9% vs 51.9%, P 5 .9; Figure 1E-G). Altogether, these results indicate that STEVOR-mediated rigidity of infected erythrocytes is dependent on the C-terminal domain in both asexual and gametocytes stages.…”

Transcriptional profiling defines dynamics of parasite tissue sequestration during malaria infection

“…It is well established that ring-infected RBCs retain deformability properties of uninfected RBCs while the presence of more mature parasites significantly increases the rigidity of the host cell [64,65]. This switch in cellular rigidity coincides with induction of adhesive properties due to PfEMP1 surface display at around 20 hours post-invasion [66,67]. Transcriptional activity of the PfEMP1-encoding var genes is rapidly down-regulated once clinical isolates are cultured ex vivo [28], and such transcriptional and physiological changes of the parasite during culture adaptation are of concern when conducting in vitro studies.…”

Spleen-Dependent Regulation of Antigenic Variation in Malaria Parasites: Plasmodium knowlesi SICAvar Expression Profiles in Splenic and Asplenic Hosts

“…While clearly challenging for P. falciparum studies, the importance of studying antigenic variation in the context of the host environment has been stressed [1,66] and an increasing number of studies over the last decade have aimed to compare, contrast, and model results generated from in vivo or ex vivo iRBC samples acquired from human P. falciparum infections [26,59,66-75]. Such studies are also beginning to recognize the potential impact on var gene expression and switching depending on whether individuals are naïve and experiencing a more severe case of malaria or have previous exposure to the parasite and some degree of adaptive immunity.…”