Abstract
ABSTRACTPlasmodium 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.
Highlights
Plasmodium falciparum (Pf) malaria parasites remodel their host human erythrocytes to establish an environment suitable for their growth and replication
The newly formed parasitophorous vacuole membrane (PVM) is continuous with host erythrocyte but some erythrocyte proteins are excluded from PVM (Ward et al, 1993) by complex molecular events that occur at the membrane junction that forms during parasite invasion (Murphy et al, 2004; Cowman et al, 2012), suggesting that molecular organization of the PVM and parasitized erythrocyte membrane (EM) differ
To study the properties of membranes and cholesterol distributions in the pRBC and non-parasitized erythrocytes, we used a membrane environment-sensitive fluorophore (1-[2-Hydroxy-3-(N,N-di-methylN-hydroxyethyl)ammoniopropyl]-4-[b-[2-(di-n-butylamino)-6-napthyl] vinyl]pyridinium dibromide (Di-4 ANEPPDHQ or ‘‘Di-4’’)) (Obaid et al, 2004), which shows a blue-shift in emission spectrum of as much as 60 nm when it is inside a cholesterol-rich membrane phase (Jin et al, 2005)
Summary
Plasmodium falciparum (Pf) malaria parasites remodel their host human erythrocytes to establish an environment suitable for their growth and replication. This remodeled environment includes the single bilayer membrane system of a parasitophorous. A translocon complex termed PTEX recognizes export motifs (Marti et al, 2004) of many parasite proteins destined for host erythrocyte cytoplasm and membrane (de Koning-Ward et al, 2009); much remains unknown of the steps by which the PVM, TVN and MC develop and how shapes and biophysical properties of their membranes support trafficking of proteins and other molecules
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