scholarly journals Plasmodium falciparum-infected erythrocytes utilize a synthetic truncated ceramide precursor for synthesis and secretion of truncated sphingomyelin

1995 ◽  
Vol 308 (1) ◽  
pp. 335-341 ◽  
Author(s):  
I Ansorge ◽  
D Jeckel ◽  
F Wieland ◽  
K Lingelbach
Keyword(s):  
Plasmodium Falciparum ◽  
Mammalian Cells ◽  
Membrane Fraction ◽  
Infected Erythrocyte ◽  
Brefeldin A ◽  
Parasite Development ◽  
Phospholipid Content ◽  
Unicellular Eukaryote ◽  
Fungal Metabolite ◽  
Sphingomyelin Synthase

Plasmodium falciparum is an intracellular parasite of human erythrocytes. Parasite development is accompanied by an increase of the phospholipid content of the infected erythrocyte, but it results in a selective decrease of sphingomyelin. We have studied sphingomyelin biosynthesis in infected erythrocytes using as substrate a synthetic radiolabelled ceramide precursor, truncated in both hydrophobic chains. Lysates of infected, unlike those of non-infected, erythrocytes contained sphingomyelin synthase activity, which therefore is of parasite origin. The enzyme activity was associated with a membrane fraction. In contrast to mammalian cells, the parasite did not synthesize detectable levels of glycosphingolipids. In intact infected erythrocytes the ceramide precursor was converted into a correspondingly truncated soluble sphingomyelin which was released into the medium at 37 degrees C. Release of truncated sphingomyelin was inhibited by low temperature (15 degrees C) but not by the fungal metabolite brefeldin A which, however, arrests protein export from the parasite. While membranes of mammalian cells, including the plasma membrane of non-infected erythrocytes, are impermeable to truncated sphingomyelin, the membrane of infected erythrocytes allowed passage of the molecule in both directions. The results obtained with the unicellular eukaryote used here as an experimental model are discussed in comparison with sphingomyelin synthesis and transport in mammalian cells.

scholarly journals Biosynthesis, export and processing of a 45 kDa protein detected in membrane clefts of erythrocytes infected with Plasmodium falciparum

1994 ◽  
Vol 302 (2) ◽  
pp. 487-496 ◽  
Author(s):  
A Das ◽  
H G Elmendorf ◽  
W I Li ◽  
K Haldar
Keyword(s):  
Plasmodium Falciparum ◽  
Life Cycle ◽  
Mammalian Cells ◽  
Secretory Pathway ◽  
Brefeldin A ◽  
Soluble Form ◽  
Parasite Development ◽  
Parasite Protein ◽  
Parasite Plasmodium Falciparum ◽  
Membrane Bound

During its asexual life cycle, the human malaria parasite Plasmodium falciparum exports numerous proteins beyond its surface to its host erythrocyte. We have studied the biosynthesis, processing and export of a 45 kDa parasite protein resident in membrane clefts in the erythrocyte cytoplasm. Our results indicate that this cleft protein is made as a single tightly membrane-bound 45 kDa polypeptide in ring- and trophozoite-infected erythrocytes (0-36 h in the life cycle). Using ring/trophozoite parasites released from erythrocytes, the 45 kDa protein is shown to be efficiently transported to the cell surface. This export is specifically blocked by the drug brefeldin A, and at 15 and 20 degrees C. These results indicate that transport blocks seen in the Golgi of mammalian cells are conserved in P. falciparum. Further, the newly synthesized 45 kDa protein passes through parasite Golgi compartments before its export to clefts in the erythrocyte. In mid-to-late-ring-infected erythrocytes, a fraction of the newly synthesized 45 kDa protein is processed to a second membrane-bound phosphorylated 47 kDa protein. The t1/2 of this processing step is about 4 h, suggesting that it occurs subsequent to protein export from the parasite. Evidence is presented that, in later trophozoite stages (24-36 h), the exported 45 and 47 kDa proteins are partially converted into soluble molecules in the intraerythrocytic space. Taken together, the results indicate that the lower eukaryote P. falciparum modulates a classical secretory pathway to support membrane export beyond its plasma membrane to clefts in the erythrocyte. Subsequent to export, phosphorylation and/or conversion into a soluble form may regulate the interactions of the 45 kDa protein with the clefts during parasite development.

scholarly journals Trafficking and Association of Plasmodium falciparum MC-2TM with the Maurer’s Clefts

Pathogens ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 431
Author(s):  
Raghavendra Yadavalli ◽  
John W. Peterson ◽  
Judith A. Drazba ◽  
Tobili Y. Sam-Yellowe
Keyword(s):  
Plasmodium Falciparum ◽  
Erythrocyte Membrane ◽  
Sodium Carbonate ◽  
Infected Erythrocyte ◽  
Brefeldin A ◽  
Specific Expression ◽  
Lipid Interactions ◽  
And Topology ◽  
Streptolysin O ◽  
Maurer's Clefts

In this study, we investigated stage specific expression, trafficking, solubility and topology of endogenous PfMC-2TM in P. falciparum (3D7) infected erythrocytes. Following Brefeldin A (BFA) treatment of parasites, PfMC-2TM traffic was evaluated using immunofluorescence with antibodies reactive with PfMC-2TM. PfMC-2TM is sensitive to BFA treatment and permeabilization of infected erythrocytes with streptolysin O (SLO) and saponin, showed that the N and C-termini of PfMC-2TM are exposed to the erythrocyte cytoplasm with the central portion of the protein protected in the MC membranes. PfMC-2TM was expressed as early as 4 h post invasion (hpi), was tightly colocalized with REX-1 and trafficked to the erythrocyte membrane without a change in solubility. PfMC-2TM associated with the MC and infected erythrocyte membrane and was resistant to extraction with alkaline sodium carbonate, suggestive of protein-lipid interactions with membranes of the MC and erythrocyte. PfMC-2TM is an additional marker of the nascent MCs.

scholarly journals Inhibition of protein N-myristoylation blocks Plasmodium falciparum intraerythrocytic development, egress and invasion

PLoS Biology ◽  
2021 ◽  
Vol 19 (10) ◽  
pp. e3001408
Author(s):  
Anja C. Schlott ◽  
Ellen Knuepfer ◽  
Judith L. Green ◽  
Philip Hobson ◽  
Aaron J. Borg ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Infected Erythrocyte ◽  
Complex Function ◽  
Pleiotropic Effect ◽  
Substantial Effect ◽  
Parasite Development ◽  
Human Malaria Parasite ◽  
Erythrocyte Invasion ◽  
Erythrocytic Cycle ◽  
Parasite Egress

We have combined chemical biology and genetic modification approaches to investigate the importance of protein myristoylation in the human malaria parasite, Plasmodium falciparum. Parasite treatment during schizogony in the last 10 to 15 hours of the erythrocytic cycle with IMP-1002, an inhibitor of N-myristoyl transferase (NMT), led to a significant blockade in parasite egress from the infected erythrocyte. Two rhoptry proteins were mislocalized in the cell, suggesting that rhoptry function is disrupted. We identified 16 NMT substrates for which myristoylation was significantly reduced by NMT inhibitor (NMTi) treatment, and, of these, 6 proteins were substantially reduced in abundance. In a viability screen, we showed that for 4 of these proteins replacement of the N-terminal glycine with alanine to prevent myristoylation had a substantial effect on parasite fitness. In detailed studies of one NMT substrate, glideosome-associated protein 45 (GAP45), loss of myristoylation had no impact on protein location or glideosome assembly, in contrast to the disruption caused by GAP45 gene deletion, but GAP45 myristoylation was essential for erythrocyte invasion. Therefore, there are at least 3 mechanisms by which inhibition of NMT can disrupt parasite development and growth: early in parasite development, leading to the inhibition of schizogony and formation of “pseudoschizonts,” which has been described previously; at the end of schizogony, with disruption of rhoptry formation, merozoite development and egress from the infected erythrocyte; and at invasion, when impairment of motor complex function prevents invasion of new erythrocytes. These results underline the importance of P. falciparum NMT as a drug target because of the pleiotropic effect of its inhibition.

scholarly journals Plasmodium falciparum exports the Golgi marker sphingomyelin synthase into a tubovesicular network in the cytoplasm of mature erythrocytes

1994 ◽  
Vol 124 (4) ◽  
pp. 449-462 ◽  
Author(s):  
HG Elmendorf ◽  
K Haldar
Keyword(s):  
Plasmodium Falciparum ◽  
Plasma Membrane ◽  
Mammalian Cells ◽  
Three Dimensional ◽  
Eukaryotic Cell ◽  
Biosynthetic Activity ◽  
Trophozoite Stage ◽  
Mature Erythrocyte ◽  
Sphingomyelin Synthase ◽  
Membrane Development

This work describes two unusual features of membrane development in a eukaryotic cell. (a) The induction of an extensive network of tubovesicular membranes by the malaria parasite Plasmodium falciparum in the cytoplasm of the mature erythrocyte, and its visualization with two ceramide analogues C5-DMB-ceramide and C6-NBD-ceramide. "Sectioning" of the infected erythrocytes using laser confocal microscopy has allowed the reconstruction of detailed three-dimensional images of this novel membrane network. (b) The stage-specific export of sphingomyelin synthase, a biosynthetic activity concentrated in the Golgi of mammalian cells, to this tubovesicular network. Evidence is presented that in the extracellular merozoite stage the parasite retains sphingomyelin synthase within its plasma membrane. However, intracellular ring- and trophozoite-stage parasites export a substantial fraction (approximately 26%) of sphingomyelin synthase activity to membranes beyond their plasma membrane. Importantly we do not observe synthesis of new enzyme during these intracellular stages. Taken together these results strongly suggest that the export of this classic Golgi enzyme is developmentally regulated in Plasmodium. We discuss the significance of this export and the tubovesicular network with respect to membrane development and function in the erythrocyte cytosol.

scholarly journals Variable oxygen environments and DNMT2 determine the DNA cytosine epigenetic landscape of Plasmodium falciparum

2021 ◽  
Author(s):  
Artur Scherf ◽  
Elie Hammam ◽  
Samia Miled ◽  
Frederic Bonhomme ◽  
Benoit Arcangioli ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Mammalian Cells ◽  
Dna Methyltransferase ◽  
Cytosine Methylation ◽  
Parasite Development ◽  
Epigenetic Landscape ◽  
Human Malaria Parasite ◽  
Parasite Plasmodium ◽  
Parasite Plasmodium Falciparum ◽  
Oxidized Products

DNA cytosine methylation and its oxidized products are important epigenetic modifications in mammalian cells. Although 5-methylcytosine (5mC) was detected in the human malaria parasite Plasmodium falciparum, the presence of oxidized 5mC forms remain to be characterized.Here we establish a protocol to explore nuclease-based DNA digestion for the extremely AT-rich genome of P. falciparum (>80% A+T) for quantitative LC-MS/MS analysis of 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). We demonstrate the presence of 5hmC, 5fC and 5caC cytosine modifications in a DNMT2-only organism and observe striking ratio changes between 5mC and 5hmC during the 48-hour blood stage parasite development. Parasite-infected red blood cells cultured in different physiological oxygen concentrations revealed a shift in the cytosine modifications distribution towards the oxidized 5hmC and 5caC forms. In the absence of the canonical C5-DNA methyltransferase (DNMT1 and DNMT3A/B) in P. falciparum, we show that all cytosine modifications depend on the presence of DNMT2. We conclude that DNMT2 and oxygen levels are critical determinants that shape the dynamic cytosine epigenetic landscape in this human pathogen.

scholarly journals Inhibition of protein N-myristoylation blocks Plasmodium falciparum intraerythrocytic development, egress, and invasion

2020 ◽  
Author(s):  
Anja C. Schlott ◽  
Ellen Knuepfer ◽  
Judith L. Green ◽  
Philip Hobson ◽  
Aaron J. Borg ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Infected Erythrocyte ◽  
Complex Function ◽  
Pleiotropic Effect ◽  
Substantial Effect ◽  
Parasite Development ◽  
Human Malaria Parasite ◽  
Erythrocyte Invasion ◽  
Erythrocytic Cycle ◽  
Parasite Egress

ABSTRACTWe have combined chemical biology and genetic modification approaches to investigate the importance of protein myristoylation in the human malaria parasite, Plasmodium falciparum. Parasite treatment during schizogony in the last ten to fifteen hours of the erythrocytic cycle with IMP-1002, an inhibitor of N-myristoyl transferase (NMT), led to a significant blockade in parasite egress from the infected erythrocyte. Two rhoptry proteins were mislocalized in the cell, suggesting that rhoptry function is disrupted. We identified sixteen NMT substrates for which myristoylation was significantly reduced by NMT inhibitor treatment, and of these, six proteins were substantially reduced in abundance. In a viability screen, we showed that for four of these proteins replacement of the N-terminal glycine with alanine to prevent myristoylation had a substantial effect on parasite fitness. In detailed studies of one NMT substrate, glideosome associated protein 45 (GAP45), loss of myristoylation had no impact on protein location or glideosome assembly, in contrast to the disruption caused by GAP45 gene deletion, but GAP45 myristoylation was essential for erythrocyte invasion. Therefore, there are at least three mechanisms by which inhibition of NMT can disrupt parasite development and growth: early in parasite development, leading to the inhibition of schizogony and formation of ‘pseudoschizonts’, which has been described previously; at the end of schizogony, with disruption of rhoptry formation, merozoite development and egress from the infected erythrocyte; and at invasion, when impairment of motor complex function prevents invasion of new erythrocytes. These results underline the importance of P. falciparum NMT as a drug target because of the pleiotropic effect of its inhibition.

Combinatorial Metabolism in Plasmodium falciparum-Infected Erythrocyte and Interplay of Glycosylation & Phosphorylation

2004 ◽  
Vol 8 (5) ◽  
pp. 453-461 ◽  
Author(s):  
Bentham Science Publisher Nasir-ud-Din ◽  
Ishtiaq Ahmad ◽  
Asma Iqbal ◽  
Daniel Hoessli
Keyword(s):  
Plasmodium Falciparum ◽  
Infected Erythrocyte

scholarly journals Stage-dependent effect of deferoxamine on growth of Plasmodium falciparum in vitro

Blood ◽  
1990 ◽  
Vol 76 (6) ◽  
pp. 1250-1255 ◽  
Author(s):  
S Whitehead ◽  
TE Peto
Keyword(s):  
Plasmodium Falciparum ◽  
Growth Inhibition ◽  
Antimalarial Activity ◽  
Clinical Malaria ◽  
Inhibitory Effect ◽  
Parasite Development ◽  
And Control ◽  
Speed Of Onset

Abstract Deferoxamine (DF) has antimalarial activity that can be demonstrated in vitro and in vivo. This study is designed to examine the speed of onset and stage dependency of growth inhibition by DF and to determine whether its antimalarial activity is cytostatic or cytocidal. Growth inhibition was assessed by suppression of hypoxanthine incorporation and differences in morphologic appearance between treated and control parasites. Using synchronized in vitro cultures of Plasmodium falciparum, growth inhibition by DF was detected within a single parasite cycle. Ring and nonpigmented trophozoite stages were sensitive to the inhibitory effect of DF but cytostatic antimalarial activity was suggested by evidence of parasite recovery in later cycles. However, profound growth inhibition, with no evidence of subsequent recovery, occurred when pigmented trophozoites and early schizonts were exposed to DF. At this stage in parasite development, the activity of DF was cytocidal and furthermore, the critical period of exposure may be as short as 6 hours. These observations suggest that iron chelators may have a role in the treatment of clinical malaria.

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