Plasmodium falciparum erythrocyte-binding antigen 175 triggers a biophysical change in the red blood cell that facilitates invasion

Abstract

Invasion of the red blood cell (RBC) by the Plasmodium parasite defines the start of malaria disease pathogenesis. To date, experimental investigations into invasion have focused predominantly on the role of parasite adhesins or signaling pathways and the identity of binding receptors on the red cell surface. A potential role for signaling pathways within the erythrocyte, which might alter red cell biophysical properties to facilitate invasion, has largely been ignored. The parasite erythrocyte-binding antigen 175 (EBA175), a protein required for entry in most parasite strains, plays a key role by binding to glycophorin A (GPA) on the red cell surface, although the function of this binding interaction is unknown. Here, using real-time deformability cytometry and flicker spectroscopy to define biophysical properties of the erythrocyte, we show that EBA175 binding to GPA leads to an increase in the cytoskeletal tension of the red cell and a reduction in the bending modulus of the cell's membrane. We isolate the changes in the cytoskeleton and membrane and show that reduction in the bending modulus is directly correlated with parasite invasion efficiency. These data strongly imply that the malaria parasite primes the erythrocyte surface through its binding antigens, altering the biophysical nature of the target cell and thus reducing a critical energy barrier to invasion. This finding would constitute a major change in our concept of malaria parasite invasion, suggesting it is, in fact, a balance between parasite and host cell physical forces working together to facilitate entry.erythrocyte | malaria | real-time deformability cytometry | flicker spectroscopy | merozoite M alaria infections cause ∼438,000 deaths per year, most of which are due to the protozoan parasite Plasmodium falciparum (1). Although extensive eradication efforts have helped to reduce the incidence of malaria, the spread of drug resistance is a growing concern and novel treatments are urgently needed (2). Throughout the Plasmodium life cycle, parasites shuttle between replicative and motile life-cycle stages, with the motile forms or "zoites" highly adapted to the invasion of host cells. During the blood stages of infection, the merozoite targets and invades the red blood cell (RBC) rapidly (< 30 s) in a process that involves numerous parasite ligands and host cell receptors (3). Merozoite entry is a multistep process commencing with initial attachment and ending with parasite actomyosin-driven invasion (4). Although extensive cellular and molecular details of each step have been elucidated (5, 6), there is still little mechanistic understanding of the role of each protein involved or the signaling events within the erythrocyte that accompany entry (7).Initial attachment to the RBC surface is likely mediated by merozoite surface proteins. Subsequently, the key signaling and strong attachment interactions between host and parasite membranes are thought to be mediated by two major classes of adhesins released either before or concomitant with inva...

The effects of dyslipidaemia and cholesterol modulation on erythrocyte susceptibility to malaria parasite infection

“…Thus, in an attempt to increase erythrocyte cholesterol content, red cells were incubated with MβCD coupled cholesterol. As expected, a highly significant increase in erythrocyte bending modulus was observed, whilst no significant increase in membrane tension (a measure of cytoskeletal effects (Koch, et al 2017) was found at this concentration ( Fig 1E-F). To confirm that additional cholesterol had been incorporated into the cell membranes, cells were washed, lipid membrane extracted, purified, dried and resolubilised to measure cholesterol content.…”

“…Based on our previous findings demonstrating that a reduction in the erythrocyte bending modulus resulted in an increased merozoite invasion efficiency (Koch, et al 2017), we hypothesised that stiffening the erythrocyte membrane by increasing cholesterol content would result in a lower merozoite invasion efficiency. Erythrocytes were incubated in media supplemented with a range of cholesterol concentrations, however, no change in the red cell bending modulus was found ( Fig 1A-D).…”

“…There is a detailed appreciation of the stepwise molecular events that characterise merozoite invasion (Cowman, et al 2012), however, the role the erythrocyte plays in the process was, up until recently, largely overlooked (Koch and Baum 2016). There has been, however, a growing appreciation in the past few years that parasite binding to the erythrocyte, stimulates biophysical changes in the red cell that likely facilitate entry making it energetically more favourable (Koch, et al 2017, Sisquella, et al 2017. Further, several key polymorphisms that protect against malaria infection may do so directly by modulating erythrocyte biophysical properties (Genton, et al 1995, Ndila, et al 2018.…”

High-throughput microfluidic characterization of erythrocyte shape and mechanical variability

“…This can be useful in blood analysis, since RBC deformability might be altered in many blood related diseases and disorders, such as malaria, sickle-cell anemia, diabetes, etc. (13,14,21).…”

“…Changes in elastic properties and shape of RBCs are of great interest in microcirculation research, because they affect many essential microvascular processes, including blood flow resistance (3)(4)(5), the viscosity of whole blood (6)(7)(8)(9), ATP release and oxygen delivery (10)(11)(12). Alterations in RBC material properties and shape are often associated with different blood diseases and disorders (13,14), such as sickle cell anemia (15,16), diabetes mellitus (17,18), and malaria (19)(20)(21). The importance of RBC behavior in flow also motivates large research efforts in blood processing and analysis using microfluidic devices (22)(23)(24)(25)(26).…”