Inward cholesterol gradient of the membrane system in P. falciparum-infected erythrocytes involves a dilution effect from parasite-produced lipids

Abstract

Plasmodium falciparum (Pf) infection remodels the human erythrocyte with new membrane systems, including a modified host erythrocyte membrane (EM), a parasitophorous vacuole membrane (PVM), a tubulovesicular network (TVN), and Maurer's clefts (MC). Here we report on the relative cholesterol contents of these membranes in parasitized normal (HbAA) and hemoglobin S-containing (HbAS, HbAS) erythrocytes. Results from fluorescence lifetime imaging microscopy (FLIM) experiments with a cholesterol-sensitive fluorophore show that membrane cholesterol levels in parasitized erythrocytes (pRBC) decrease inwardly from the EM, to the MC/TVN, to the PVM, and finally to the parasite membrane (PM). Cholesterol depletion of pRBC by methyl-β-cyclodextrin treatment caused a collapse of this gradient. Lipid and cholesterol exchange data suggest that the cholesterol gradient involves a dilution effect from non-sterol lipids produced by the parasite. FLIM signals from the PVM or PM showed little or no difference between parasitized HbAA vs HbS-containing erythrocytes that differed in lipid content, suggesting that malaria parasites may regulate the cholesterol contents of the PVM and PM independently of levels in the host cell membrane. Cholesterol levels may affect raft structures and the membrane trafficking and sorting functions that support Pf survival in HbAA, HbAS and HbSS erythrocytes.

Recent progress on lipid lateral heterogeneity in plasma membranes: From rafts to submicrometric domains

“…The use of different beam radii for photobleaching fluorescent lipid analogs has allowed to infer the existence of submicrometric lipid domains [19, 30, 146]. Fluorescence Lifetime Imaging Microscopy (FLIM) has been used to detect submicrometric domains in Laurdan-labeled NIH 3T3 fibroblasts or upon RBC infection by Plasmodium falciparum, which creates areas of cholesterol heterogeneity [147, 148]. Fluorescence Correlation Spectroscopy (FCS) can determine molecular concentration, diffusion as well as intra- and inter-molecular interactions.…”

Imaging of the subsurface structures of “unroofed” Plasmodium falciparum-infected erythrocytes

“…New membranous systems including Maurer's clefts (MC) and the tubulovesicular network (TVN) support the trafficking and placement of parasite proteins such as the variable cytoadherence protein P. falciparum Erythrocyte Membrane Protein 1 (PfEMP1) (Elmendorf and Haldar, 1994;Tilley et al, 2008;Wickert and Krohne, 2007;Wickert et al, 2003). Various studies have reported that MC can communicate through the TVN to the parasitophorous vacuolar membrane (PVM), and that they may also connect with the host erythrocyte membrane by small vesicles associated with filaments and by stalk-like structures or ducts to the membrane of the host cell (Cyrklaff et al, 2011;Hanssen et al, 2008aHanssen et al, , 2010Pouvelle et al, 1991;Taraschi et al, 2001Taraschi et al, , 2003Tokumasu et al, 2014).…”

Plasmodium falciparum Niemann-Pick Type C1-Related Protein is a Druggable Target Required for Parasite Membrane Homeostasis

“…Cholesterol is not synthesized by malaria parasites but is taken up from the erythrocyte and incorporated into parasite membranes. An inward cholesterol gradient is formed as the parasite grows 31 . Resistance of the PPM to saponin permeation is believed to be due to a dearth of cholesterol within the PPM.…”