Plasmodium falciparum: Generation of pure gametocyte culture by heparin treatment

Miao, Jun; Wang, Zenglei; Liu, Min; Parker, Daniel M.; Li, Xiaolian; Chen, Xiaoguang

doi:10.1016/j.exppara.2013.09.010

Cited by 38 publications

(31 citation statements)

References 24 publications

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“…Gametocyte induction was as previously described with heparin sodium salt (Sigma-Aldrich) included to inhibit asexual parasite proliferation in gametocyte cultures (Miao et al., 2013). Late stage II gametocytes were purified by a 75%/35% Percoll gradient on day 4 post gametocyte induction.…”

Section: Methodsmentioning

confidence: 99%

Interactions between tafenoquine and artemisinin-combination therapy partner drug in asexual and sexual stage Plasmodium falciparum

Kemirembe

Cabrera

2017

International Journal for Parasitology: Drugs and Drug Resistan

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The 8-aminoquinoline tafenoquine (TFQ), a primaquine derivative, is currently in late-stage clinical development for the radical cure of P. vivax. Here drug interactions between TFQ and chloroquine and six artemisinin-combination therapy (ACT) partner drugs in P. falciparum asexual stages and gametocytes were investigated. TFQ was mostly synergistic with the ACT-partner drugs in asexual parasites regardless of genetic backgrounds. However, at fixed ratios of 1:3, 1:1 and 3:1, TFQ only interacted synergistically with naphthoquine, pyronaridine and piperaquine in gametocytes. This study indicated that TFQ and ACT-partner drugs will likely have increased potency against asexual stages of the malaria parasites, whereas some drugs may interfere with each other against the P. falciparum gametocytes.

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Section: Methodsmentioning

confidence: 99%

Interactions between tafenoquine and artemisinin-combination therapy partner drug in asexual and sexual stage Plasmodium falciparum

Kemirembe

Cabrera

2017

International Journal for Parasitology: Drugs and Drug Resistan

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“…Cells were treated with MitoTracker Red CMXRos dye (Invitrogen) for mitochondrial labelling at a final concentration of 50 nM in parasite culture for half an hour. Gametocytes were generated using heparin according the protocol described earlier 50 . Different blood stages of the parasite were fixed and processed for immunofluorescence studies using the protocol described earlier 51 .…”

Section: Methodsmentioning

confidence: 99%

Structural and functional attributes of malaria parasite diadenosine tetraphosphate hydrolase

Sharma

Yogavel

2016

Sci Rep

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Malaria symptoms are driven by periodic multiplication cycles of Plasmodium parasites in human red blood corpuscles (RBCs). Malaria infection still accounts for ~600,000 annual deaths, and hence discovery of both new drug targets and drugs remains vital. In the present study, we have investigated the malaria parasite enzyme diadenosine tetraphosphate (Ap4A) hydrolase that regulates levels of signalling molecules like Ap4A by hydrolyzing them to ATP and AMP. We have tracked the spatial distribution of parasitic Ap4A hydrolase in infected RBCs, and reveal its unusual localization on the infected RBC membrane in subpopulation of infected cells. Interestingly, enzyme activity assays reveal an interaction between Ap4A hydrolase and the parasite growth inhibitor suramin. We also present a high resolution crystal structure of Ap4A hydrolase in apo- and sulphate- bound state, where the sulphate resides in the enzyme active site by mimicking the phosphate of substrates like Ap4A. The unexpected infected erythrocyte localization of the parasitic Ap4A hydrolase hints at a possible role of this enzyme in purinerigic signaling. In addition, atomic structure of Ap4A hydrolase provides insights for selective drug targeting.

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“…For the isolation of cellular RNA from different developmental stages, synchronized P . falciparum in vitro cultures for each state and gametocyte cultures [ 26 ] were employed. Cells were lysed with a QIAshredder and extraction of cellular RNA followed subsequently according to the protocol of the RNAeasy Minikit (Qiagen, Hilden, Germany).…”

Section: Methodsmentioning

confidence: 99%

A Putative Non-Canonical Ras-Like GTPase from P. falciparum: Chemical Properties and Characterization of the Protein

et al. 2015

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During its development the malaria parasite P. falciparum has to adapt to various different environmental contexts. Key cellular mechanisms involving G-protein coupled signal transduction chains are assumed to act at these interfaces. Heterotrimeric G-proteins are absent in Plasmodium. We here describe the first cloning and expression of a putative, non-canonical Ras-like G protein (acronym PfG) from Plasmodium. PfG reveals an open reading frame of 2736 bp encoding a protein of 912 amino acids with a theoretical pI of 8.68 and a molecular weight of 108.57 kDa. Transcript levels and expression are significantly increased in the erythrocytic phase in particular during schizont and gametocyte formation. Most notably, PfG has GTP binding capacity and GTPase activity due to an EngA2 domain present in small Ras-like GTPases in a variety of Bacillus species and Mycobacteria. By contrast, plasmodial PfG is divergent from any human alpha-subunit. PfG was expressed in E. coli as a histidine-tagged fusion protein and was stable only for 3.5 hours. Purification was only possible under native conditions by Nickel-chelate chromatography and subsequent separation by Blue Native PAGE. Binding of a fluorescent GTP analogue BODIPY® FL guanosine 5’O-(thiotriphosphate) was determined by fluorescence emission. Mastoparan stimulated GTP binding in the presence of Mg2+. GTPase activity was determined colorimetrically. Activity expressed as absolute fluorescence was 50% higher for the human paralogue than the activity of the parasitic enzyme. The PfG protein is expressed in the erythrocytic stages and binds GTP after immunoprecipitation. Immunofluorescence using specific antiserum suggests that PfG localizes to the parasite cytosol. The current data suggest that the putitative, Ras-like G-protein might be involved in a non-canonical signaling pathway in Plasmodium. Research on the function of PfG with respect to pathogenesis and antimalarial chemotherapy is currently under way.

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Plasmodium falciparum: Generation of pure gametocyte culture by heparin treatment

Cited by 38 publications

References 24 publications

Interactions between tafenoquine and artemisinin-combination therapy partner drug in asexual and sexual stage Plasmodium falciparum

Interactions between tafenoquine and artemisinin-combination therapy partner drug in asexual and sexual stage Plasmodium falciparum

Structural and functional attributes of malaria parasite diadenosine tetraphosphate hydrolase

A Putative Non-Canonical Ras-Like GTPase from P. falciparum: Chemical Properties and Characterization of the Protein

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