A Central Role for P48/45 in Malaria Parasite Male Gamete Fertility

Laboratory for Parasitology, Leiden University Medical Center, P.O. Box 9600, 2300 RC Leiden, The Netherlands.
Cell (Impact Factor: 32.24). 02/2001; 104(1):153-64. DOI: 10.1016/S0092-8674(01)00199-4
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Fertilization and zygote development are obligate features of the malaria parasite life cycle and occur during parasite transmission to mosquitoes. The surface protein PFS48/45 is expressed by male and female gametes of Plasmodium falciparum and PFS48/45 antibodies prevent zygote development and transmission. Here, gene disruption was used to show that Pfs48/45 and the ortholog Pbs48/45 from a rodent malaria parasite P. berghei play a conserved and important role in fertilization. p48/45- parasites had a reduced capacity to produce oocysts in mosquitoes due to greatly reduced zygote formation. Unexpectedly, only male gamete fertility of p48/45- parasites was affected, failing to penetrate otherwise fertile female gametes. P48/45 is shown to be a surface protein of malaria parasites with a demonstrable role in fertilization.

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Available from: Joanna A M Braks, Mar 10, 2015
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    • "Fertilisation and development of the malaria parasite in the mosquito midgut is a crucial 56 process for malaria transmission, which in P. falciparum can be interrupted by human antibodies 57 against a number of sexual stage proteins (Ouedraogo et al., 2011), and several of these have 58 been investigated as potential malaria transmission-blocking vaccine antigens. Pfs230 and 59 Pfs48/45 belong to the 6-cysteine family and are proteins expressed on the surface of gametes 60 (Carter et al., 1995; van Dijk et al., 2001, 2010). CCP/LAP proteins are another group of antigens 61 expressed during gametocyte differentiation and gametogenesis (Delrieu et al., 2002; Pradel et al., 62 2004; Scholz et al., 2008) which, when disrupted, lead to blockage of the translation of sporozoites 63 from the oocyst on the mosquito midgut wall to the salivary gland. "
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    ABSTRACT: Plasmodium ovale curtisi and Plasmodium ovale wallikeri are distinct species of malaria parasite which are sympatric throughout the tropics, except for the Americas. Despite this complete overlap in geographic range, these two species do not recombine. Although morphologically very similar, the two taxa must possess distinct characters which prevent recombination between them. We hypothesised that proteins required for sexual reproduction have sufficiently diverged between the two species to prevent recombination in any mosquito blood meal in which gametocytes of both species are ingested. In order to investigate possible barriers to inter-species mating between P. ovale curtisi and P. ovale wallikeri, homologues of genes encoding sexual stage proteins in other plasmodia were identified and compared between the two species. Database searches with motifs for 6-cysteine, Limulus Coagulation factor C domain-containing proteins and other relevant sexual stage proteins in the genus Plasmodium were performed in the available P. ovale curtisi partial genome database (Wellcome Trust Sanger Institute, UK). Sequence fragments obtained were used as the basis for PCR walking along each gene of interest in reference isolates of both P. ovale curtisi and P. ovale wallikeri. Sequence alignment of the homologues of each gene in each species showed complete dimorphism across all isolates. In conclusion, substantial divergence between sexual stage proteins in the two P. ovale spp. was observed, providing further evidence that these do not recombine in nature. Incompatibility of proteins involved in sexual development and fertilisation thus remains a plausible explanation for the observed lack of natural recombination between P. ovale curtisi and P. ovale wallikeri. Copyright © 2015. Published by Elsevier Ltd.
    International journal for parasitology 03/2015; 158(7). DOI:10.1016/j.ijpara.2015.02.004 · 3.87 Impact Factor
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    • "Once inside the mosquito midgut the parasite emerges from the erythrocyte as a gamete, exposing Pfs48/45 on the surface where it can be targeted by antibodies and other components of the blood meal [4]. The cellular localization and the critical role of Pfs48/45 for male gamete fertility [8] make Pfs48/45 a lead candidate for clinical vaccine development. Pfs48/45 in general and the C-terminal portion in particular is relatively cysteine-rich with multiple disulfide bonds resulting in "
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    ABSTRACT: The sexual stage Pfs48/45 antigen is a well-established lead candidate for a transmission blocking (TB) vaccine because of its critical role in parasite fertilization. We have recently produced the carboxy-terminal 10C-fragment of Pfs48/45 containing three known epitopes for TB antibodies as a chimera with the N-terminal region of GLURP (R0). The resulting fusion protein elicited high titer TB antibodies in rodents. To increase the relatively low yield of correctly folded Pfs48/45 we have generated a series of novel chimera truncating the 10C-fragments to 6 cysteine residues containing sub-units (6C). All constructs harbor the major epitope I for TB antibodies. One of these sub-units (R0.6Cc), produced high yields of correctly folded conformers, which could be purified by a simple 2-step procedure. Purified R0.6Cc was stable and elicits high titer TB antibodies in rats. The yield, purity and stability of R0.6Cc allows for further clinical development. Copyright © 2015. Published by Elsevier Ltd.
    Vaccine 02/2015; 33(16). DOI:10.1016/j.vaccine.2015.02.040 · 3.62 Impact Factor
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    • "So far, we determined a ∼ 10-fold reduction in microgamete formation, yet observed a ∼ 100-fold reduced oocyst burden in cutp− parasites. To confirm a male fertility defect and test female fertility, we performed a cross-fertilization assay and quantified the resulting ookinetes, a functional read-out for gamete fertility (van Dijk et al., 2001; 2010,). As published previously, no ookinetes were detectable in cultures containing p48/45− or p47− parasites alone, whereas in co-cultures p47− male microgametes were able to productively fertilize p48/45− females and ookinetes were readily formed (Fig. 7). "
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    ABSTRACT: Homeostasis of the trace element copper is essential to all eukaryotic life. Copper serves as a cofactor in metalloenzymes and catalyzes electron transfer reactions as well as the generation of potentially toxic reactive oxygen species. Here, we describe the functional characterization of an evolutionarily highly conserved, predicted copper-transporting P-type ATPase (CuTP) in the murine malaria model parasite Plasmodium berghei. Live imaging of a parasite line expressing a fluorescently tagged CuTP demonstrated that CuTP is predominantly located in vesicular bodies of the parasite. A P. berghei loss-of-function mutant line was readily obtained and showed no apparent defect in in vivo blood stage growth. Parasite transmission through the mosquito vector was severely affected, but not entirely abolished. We show that male and female gametocytes are abundant in cutp(-) parasites, but activation of male microgametes and exflagellation were strongly impaired. This specific defect could be mimicked by addition of the copper chelator neocuproine to wild-type gametocytes. A cross-fertilization assay demonstrated that female fertility was also severely abrogated. In conclusion, we provide experimental genetic and pharmacological evidence that a healthy copper homeostasis is critical to malaria parasite fertility of both genders of gametocyte and, hence, to transmission to the mosquito vector.
    Molecular Microbiology 11/2013; 91(2). DOI:10.1111/mmi.12461 · 4.42 Impact Factor
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