Adhesion between P. falciparum infected erythrocytes and human endothelial receptors follows alternative binding dynamics under flow and febrile conditions

Abstract

characterizing the adhesive dynamics of Plasmodium falciparum infected erythrocytes (IEs) to different endothelial cell receptors (ECRs) in flow is a big challenge considering available methods. This study investigated the adhesive dynamics of IEs to five ECRs (CD36, ICAM-1, P-selectin, CD9, CSA) using simulations of in vivo-like flow and febrile conditions. To characterize the interactions between ECRs and knobby and knobless IEs of two laboratory-adapted P. falciplarum isolates, cytoadhesion analysis over time was performed using a new tracking bioinformatics method. The results revealed that IEs performed rolling adhesion exclusively over CD36, but exhibited stationary binding to the other four ECRs. The absence of knobs affected rolling adhesion both with respect to the distance travelled by IEs and their velocity. Knobs played a critical role at febrile temperatures by stabilizing the binding interaction. Our results clearly underline the complexity of the IE-receptor interaction and the importance of knobs for the survival of the parasite at fever temperatures, and lead us to propose a new hypothesis that could open up new strategies for the treatment of malaria. Cytoadhesion of Plasmodium falciparum to human endothelial cell receptors (ECRs) causes complications and deaths following malaria infection. In 2018, 405,000 malaria-related deaths were registered (61% of which were of children younger than 5 years old) 1. Cytoadhesion leads to accumulation of infected erythrocytes (IEs) within the microvascular bed of vital organs such as the brain, lungs, and kidneys. Death can eventually occur due to decreased blood supply and organ failure 2,3. Cytoadhesion results from interactions between members of the P. falciparum erythrocyte membrane protein 1 (PfEMP1) family and different ECRs 4-8. About 60 var genes per parasite genome encode PfEMP1 family members. Only one PfEMP1 variant is located on the membrane of IEs at the trophozoite stage, but the corresponding var gene is already expressed at the ring stage in a mutually exclusive pattern 9. The most studied interaction partners are the ECRs CD36, intracellular adhesion molecule 1 (ICAM-1), endothelial protein C receptor, and chondroitin sulfate A (CSA) 6,7. In general, PfEMP1 molecules cluster on nanoscale protrusions, called knobs, located on the membrane of IEs. Knobs consist of various submembranous structural proteins, predominantly knob-associated histidine-rich protein (KAHRP) 10. KAHRP contains several binding domains that interact with both parasite and host factors. Knobs begin to appear on the surface of IEs at 16 h post-invasion (hpi). The density of knobs increases from 20 to 60/µm 2 with parasite development from the

PerTurboID: A targeted in situ method to measure changes in a local protein environment reveals the impact of kinase deletion on cytoadhesion in malaria causing parasites

“…Since Var2CSA is one of∼60 PfEMP1 variants, with only one expressed in a parasite at any given time, we wanted to test whether FIKK4.1 is important for trafficking other PfEMP1 variants. To do so, we selected for binding to other endothelial cell receptors by panning over CD36-expressing CHO cells (49). Treatment of these FIKK4.1 cKO parasites with rapamycin results in a 51.5 ± 13% reduction in cytoadhesion under flow to CHO cells expressing CD36 (Figure 5C).…”

“…Cytoadhesion assay EPCR/ICAM1-binding iRBCs were obtained via magnetic enrichment of dual-binding iRBCs with ProteinA-coupled Dynabeads (Invitrogen), which were associated to anti-EPCR/ICAM1-binding PfEMP1 antibodies (a kind gift from Anja Jensen, University of Copenhagen) as described previously (64). Late trophozoites were selected for CD36 binding by incubating percoll-enriched iRBCs with CD36-CHO cells for 1 h at 37 °C as previously described (49). Adherent cells were kept in culture after 5 x washing with RPMI.…”

Gene expression profiling of malaria parasites reveals common virulence gene expression in adult first-time infected patients and severe cases

“…Based on this and our findings we hypothesize that parasites circulating in severely ill patients are younger due to the expression of EPCR-/ICAM-1-binding Pf EMP1 variants, whereas parasites in non-severe patients may circulate longer due to their expression of Pf EMP1 binding to CD36. Although affinity of the CIDR domains to each of these receptors was shown to be similar with median Kd values of 12 nM for CD36 (Hsieh et al, 2016) and 16.5 nM for EPCR (Lau et al, 2015), recent papers describe a rolling binding phenotype of infected red blood cells over CD36 and a static binding for EPCR and ICAM-1 under flow conditions (Lubiana et al, 2020; Bernabeu et al, 2019; Dasanna et al, 2017; Herricks et al, 2013). Due to their biconcave shape trophozoites seem to roll faster but less stable by flipping over CD36-expressing cells, whereas schizonts roll over longer distances at different shear stresses applied.…”

Generation and Export of Red Blood Cell ATP in Health and Disease

“…Similarly, "knobby" (cellular-protrusionbearing) erythrocytes infected with Plasmodium falciparum (an organism frequently responsible for malarial infection) displayed greater temperature-dependent binding to ECs than "knobless" erythrocytes infected with the same pathogen. Rather than a simple physical advantage in cell-cell binding, the proadhesive effect of the presence of knobs may reflect a localized abundance of adhesive ligands and receptors (Lubiana et al, 2020). Interestingly, Leal Denis and coworkers demonstrated that the export of ATP by RBCs made to adhere to coverslips coated with poly-D-lysine was far greater than that of RBCs on uncoated coverslips (Leal Denis et al, 2013).…”