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A multidomain adhesion protein family expressed in Plasmodium falciparum is essential for transmission to the mosquito.

Pradel G, Hayton K, Aravind L, Iyer LM, Abrahamsen MS, Bonawitz A, Mejia C, Templeton TJ - J. Exp. Med. (2004)

Bottom Line: Three of these identified genes, named PfCCp1, PfCCp2, and PfCCp3, have multiple adhesive modules including a common Limulus coagulation factor C domain also found in two additional Plasmodium genes.PfCCp expression markedly decreased after formation of zygotes.Our results describe members of a conserved apicomplexan protein family expressed in sexual stage Plasmodium parasites that may represent candidates for subunits of a transmission-blocking vaccine.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Weill Medical College of Cornell University, 1300 York Avenue, New York, NY 10021, USA.

ABSTRACT
The recent sequencing of several apicomplexan genomes has provided the opportunity to characterize novel antigens essential for the parasite life cycle that might lead to the development of new diagnostic and therapeutic markers. Here we have screened the Plasmodium falciparum genome sequence for genes encoding extracellular multidomain putative adhesive proteins. Three of these identified genes, named PfCCp1, PfCCp2, and PfCCp3, have multiple adhesive modules including a common Limulus coagulation factor C domain also found in two additional Plasmodium genes. Orthologues were identified in the Cryptosporidium parvum genome sequence, indicating an evolutionary conserved function. Transcript and protein expression analysis shows sexual stage-specific expression of PfCCp1, PfCCp2, and PfCCp3, and cellular localization studies revealed plasma membrane-associated expression in mature gametocytes. During gametogenesis, PfCCps are released and localize surrounding complexes of newly emerged microgametes and macrogametes. PfCCp expression markedly decreased after formation of zygotes. To begin to address PfCCp function, the PfCCp2 and PfCCp3 gene loci were disrupted by homologous recombination, resulting in parasites capable of forming oocyst sporozoites but blocked in the salivary gland transition. Our results describe members of a conserved apicomplexan protein family expressed in sexual stage Plasmodium parasites that may represent candidates for subunits of a transmission-blocking vaccine.

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Development of midgut oocysts and sporozoites after mosquito membrane feeding with gene-disrupted parasites. Ultrastructural examinations 12 d after feeding show oocysts in mosquito midguts after feeding PfCCp2-KO clone D-11H (A) and PfCCp3-KO clone I-9C (B) cultures. Mature oocysts exhibit fully developed sporozoites for PfCCp2-KO (C) and PfCCp3-KO (D). Bars, 1 μm.
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fig8: Development of midgut oocysts and sporozoites after mosquito membrane feeding with gene-disrupted parasites. Ultrastructural examinations 12 d after feeding show oocysts in mosquito midguts after feeding PfCCp2-KO clone D-11H (A) and PfCCp3-KO clone I-9C (B) cultures. Mature oocysts exhibit fully developed sporozoites for PfCCp2-KO (C) and PfCCp3-KO (D). Bars, 1 μm.

Mentions: To characterize PfCCp2-KO and PfCCp3-KO phenotypes during mosquito transmission, mature gametocyte cultures of WT parasites and gene-disruptant clones were fed to An. freeborni mosquitoes via membrane feeding. Six independent feeding experiments for PfCCp2-KO cultures and two independent feeds for PfCCp3-KO cultures revealed a normal infection rate of mosquito midgut oocysts compared with mosquitoes fed with WT parasites (Table I). The number of oocysts per midgut was comparable in WT and gene-disruptant lines. Ultrastructural examinations of PfCCp2-KO (Fig. 8 A) and PfCCp3-KO (Fig. 8 B) oocysts indicated apparent normal differentiation, in which developed sporozoites (Fig. 8, C and D) showed no morphological differences compared with WT midgut sporozoites. However, no sporozoites were detected 19 d after feeding within or associated with salivary glands, nor within the hemocoel of mosquitoes fed with the KO lines, whereas mosquitoes fed with WT cultures exhibited salivary gland infection rates ranging from 13 to 86% (Table I). To address the fate of sporozoites, in one experiment, >50 mosquito midguts were pooled 20 d after feeding with PfCCp2-KO line F-1D, homogenized, and enriched for sporozoites via differential centrifugation. Specimens were screened by IFA using the mAb 2A10 recognizing the CS protein and revealed CP protein expression in sparse, intact midgut PfCCp2-KO sporozoites having CS protein precipitate trails indicative of motility on the glass slides (unpublished data). Thus, PfCCp2- and PfCCp3-targeted gene disruption results in a developmental blockage in the sporozoite transition from the mosquito midgut to salivary gland sporozoites.


A multidomain adhesion protein family expressed in Plasmodium falciparum is essential for transmission to the mosquito.

Pradel G, Hayton K, Aravind L, Iyer LM, Abrahamsen MS, Bonawitz A, Mejia C, Templeton TJ - J. Exp. Med. (2004)

Development of midgut oocysts and sporozoites after mosquito membrane feeding with gene-disrupted parasites. Ultrastructural examinations 12 d after feeding show oocysts in mosquito midguts after feeding PfCCp2-KO clone D-11H (A) and PfCCp3-KO clone I-9C (B) cultures. Mature oocysts exhibit fully developed sporozoites for PfCCp2-KO (C) and PfCCp3-KO (D). Bars, 1 μm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2211786&req=5

fig8: Development of midgut oocysts and sporozoites after mosquito membrane feeding with gene-disrupted parasites. Ultrastructural examinations 12 d after feeding show oocysts in mosquito midguts after feeding PfCCp2-KO clone D-11H (A) and PfCCp3-KO clone I-9C (B) cultures. Mature oocysts exhibit fully developed sporozoites for PfCCp2-KO (C) and PfCCp3-KO (D). Bars, 1 μm.
Mentions: To characterize PfCCp2-KO and PfCCp3-KO phenotypes during mosquito transmission, mature gametocyte cultures of WT parasites and gene-disruptant clones were fed to An. freeborni mosquitoes via membrane feeding. Six independent feeding experiments for PfCCp2-KO cultures and two independent feeds for PfCCp3-KO cultures revealed a normal infection rate of mosquito midgut oocysts compared with mosquitoes fed with WT parasites (Table I). The number of oocysts per midgut was comparable in WT and gene-disruptant lines. Ultrastructural examinations of PfCCp2-KO (Fig. 8 A) and PfCCp3-KO (Fig. 8 B) oocysts indicated apparent normal differentiation, in which developed sporozoites (Fig. 8, C and D) showed no morphological differences compared with WT midgut sporozoites. However, no sporozoites were detected 19 d after feeding within or associated with salivary glands, nor within the hemocoel of mosquitoes fed with the KO lines, whereas mosquitoes fed with WT cultures exhibited salivary gland infection rates ranging from 13 to 86% (Table I). To address the fate of sporozoites, in one experiment, >50 mosquito midguts were pooled 20 d after feeding with PfCCp2-KO line F-1D, homogenized, and enriched for sporozoites via differential centrifugation. Specimens were screened by IFA using the mAb 2A10 recognizing the CS protein and revealed CP protein expression in sparse, intact midgut PfCCp2-KO sporozoites having CS protein precipitate trails indicative of motility on the glass slides (unpublished data). Thus, PfCCp2- and PfCCp3-targeted gene disruption results in a developmental blockage in the sporozoite transition from the mosquito midgut to salivary gland sporozoites.

Bottom Line: Three of these identified genes, named PfCCp1, PfCCp2, and PfCCp3, have multiple adhesive modules including a common Limulus coagulation factor C domain also found in two additional Plasmodium genes.PfCCp expression markedly decreased after formation of zygotes.Our results describe members of a conserved apicomplexan protein family expressed in sexual stage Plasmodium parasites that may represent candidates for subunits of a transmission-blocking vaccine.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Weill Medical College of Cornell University, 1300 York Avenue, New York, NY 10021, USA.

ABSTRACT
The recent sequencing of several apicomplexan genomes has provided the opportunity to characterize novel antigens essential for the parasite life cycle that might lead to the development of new diagnostic and therapeutic markers. Here we have screened the Plasmodium falciparum genome sequence for genes encoding extracellular multidomain putative adhesive proteins. Three of these identified genes, named PfCCp1, PfCCp2, and PfCCp3, have multiple adhesive modules including a common Limulus coagulation factor C domain also found in two additional Plasmodium genes. Orthologues were identified in the Cryptosporidium parvum genome sequence, indicating an evolutionary conserved function. Transcript and protein expression analysis shows sexual stage-specific expression of PfCCp1, PfCCp2, and PfCCp3, and cellular localization studies revealed plasma membrane-associated expression in mature gametocytes. During gametogenesis, PfCCps are released and localize surrounding complexes of newly emerged microgametes and macrogametes. PfCCp expression markedly decreased after formation of zygotes. To begin to address PfCCp function, the PfCCp2 and PfCCp3 gene loci were disrupted by homologous recombination, resulting in parasites capable of forming oocyst sporozoites but blocked in the salivary gland transition. Our results describe members of a conserved apicomplexan protein family expressed in sexual stage Plasmodium parasites that may represent candidates for subunits of a transmission-blocking vaccine.

Show MeSH
Related in: MedlinePlus