Identification and characterisation of proteins associated with the rhoptry organelles of Plasmodium falciparum merozoites

Clark, Jessica; Anand, Rita; Akoğlu, T; McBride, Jana S.

doi:10.1007/bf00538200

Cited by 55 publications

(35 citation statements)

References 30 publications

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“…As RAP2 and RAP3 have similar molecular masses and they both interact with RAP1 (7,11,29,43), it was considered likely that they may share some homology. To test this hypothesis, the RAP2 gene sequence (42) was used to search the P. falciparum genome sequence database (http:// www.sanger.ac.uk).…”

Section: Resultsmentioning

confidence: 99%

“…Several studies established that the largest product, reported to be 86 kDa, is a short-lived precursor of the 82-kDa RAP1 protein and that this 82-kDa protein is further processed to a 67-kDa protein (7,26,27,29). The 42-kDa protein (also reported as 39 kDa) was found to be the product of a second gene, RAP2 (7,11,28,39,42). Debate remained regarding the origin of the 40-kDa (or 37-kDa) protein also immunoprecipitated as a part of the RAP1 complex.…”

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confidence: 99%

“…This complex consists of various polypeptides, ranging in size from 86 to 37 kDa, that are immunoprecipitated by antibodies to rhoptry-associated protein 1 (RAP1) (7,11,29,43). Several studies established that the largest product, reported to be 86 kDa, is a short-lived precursor of the 82-kDa RAP1 protein and that this 82-kDa protein is further processed to a 67-kDa protein (7,26,27,29).…”

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Identification and Disruption of the Gene Encoding the Third Member of the Low-Molecular-Mass Rhoptry Complex in Plasmodium falciparum

et al. 2002

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The low-molecular-mass rhoptry complex of Plasmodium falciparum consists of three proteins, rhoptryassociated protein 1 (RAP1), RAP2, and RAP3. The genes encoding RAP1 and RAP2 are known; however, the RAP3 gene has not been identified. In this study we identify the RAP3 gene from the P. falciparum genome database and show that this protein is part of the low-molecular-mass rhoptry complex. Disruption of RAP3 demonstrated that it is not essential for merozoite invasion, probably because RAP2 can complement the loss of RAP3. RAP3 has homology with RAP2, and the genes are encoded on chromosome 5 in a head-to-tail fashion. Analysis of the genome databases has identified homologous genes in all Plasmodium spp., suggesting that this protein plays a role in merozoite invasion. The region surrounding the RAP3 homologue in the Plasmodium yoelii genome is syntenic with the same region in P. falciparum; however, there is a single gene. Phylogenetic comparison of the RAP2/3 protein family from Plasmodium spp. suggests that the RAP2/3 duplication occurred after divergence of these parasite species.The merozoite form of the asexual life cycle in the blood stage of Plasmodium spp. attaches to the surface of the erythrocyte, thus initiating the invasion process (23). Ultrastructural and biochemical studies of merozoites have suggested that the rhoptries play an essential role in the invasion process. These electron-dense organelles, part of the apical complex, are connected to the surface of the apical end of the merozoite by a duct-like structure, and their contents are expelled during erythrocyte invasion (1). Proteins located on the surface of merozoites and those released from the apical organelles, the rhoptries and micronemes, are exposed to the immune system. Consequently, these proteins are potential vaccine candidates, and there is considerable interest in the identification and characterization of proteins in these organelles (2).A number of proteins located in the rhoptries have been identified and characterized, such as the low-molecular-mass rhoptry protein complex of Plasmodium falciparum (28). This complex consists of various polypeptides, ranging in size from 86 to 37 kDa, that are immunoprecipitated by antibodies to rhoptry-associated protein 1 (RAP1) (7,11,29,43). Several studies established that the largest product, reported to be 86 kDa, is a short-lived precursor of the 82-kDa RAP1 protein and that this 82-kDa protein is further processed to a 67-kDa protein (7,26,27,29). The 42-kDa protein (also reported as 39 kDa) was found to be the product of a second gene, RAP2 (7,11,28,39,42). Debate remained regarding the origin of the 40-kDa (or 37-kDa) protein also immunoprecipitated as a part of the RAP1 complex. It was suggested that this protein was a breakdown product of either RAP1 or RAP2 (41). It was subsequently shown, however, that RAP1, RAP2, and RAP3 yield different V8 protease cleavage patterns and are thus products of three distinct genes (28). The RAP2 and RAP3 proteins, in particular, were hypothesiz...

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Section: Resultsmentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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Identification and Disruption of the Gene Encoding the Third Member of the Low-Molecular-Mass Rhoptry Complex in Plasmodium falciparum

et al. 2002

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“…Subsequent studies have borne out this potential. Monoclonal antibodies (MAbs) directed against rhoptry-associated proteins have been shown to provide substantial inhibition of parasite invasion in vitro (7,17,19,22,29,37). In addition, protein preparations containing both RAP1 and RAP2 have been used to immunize Saimiri monkeys.…”

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confidence: 99%

Interaction of HLA and Age on Levels of Antibody toPlasmodium falciparumRhoptry-Associated Proteins 1 and 2

Johnson¹,

Leke

Harun

et al. 2000

Infect Immun

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The Plasmodium falciparum rhoptry-associated proteins 1 and 2 (RAP1 and RAP2) are candidate antigens for a subunit malaria vaccine. The design of the study, which looks at the acquisition of immunity to malaria from childhood to old age, has allowed us to document the interaction of HLA and age on levels of antibody to specific malarial antigens. Antibodies reach maximum levels to RAP1 after the age of 15 but to RAP2 only after the age of 30. The effect of HLA-DRB1 and -DQB1 and age on levels of antibody to rRAP1 and rRAP2 was analyzed with a multiple regression model in which all HLA alleles and age were independent variables. DQB1*0301 and -*03032 showed an age-dependent association with levels of antibody to rRAP1, being significant in children 5 to 15 years (P < 0.001) but not in individuals over 15 years of age. DRB1*03011 showed an age-dependent association with antibody levels to rRAP2; however, this association was in adults over the age of 30 years (P < 0.01) but not in individuals under the age of 30 years. No associations were detected between DRB1 alleles and RAP1 antibody levels or between DQB1 alleles and RAP2 antibody levels. Thus, not only the HLA allele but also the age at which an interaction is manifested varies for different malarial antigens. The interaction may influence either the rate of acquisition of antibody or the final level of antibody acquired by adults.The rhoptry-associated proteins 1 and 2 (RAP1 and RAP2) form a low-molecular-mass complex located in the rhoptries of Plasmodium falciparum (5, 9). The rhoptries are a pair of organelles at the apical end of the parasite that are involved in the invasion of erythrocytes; thus, molecules in the rhoptries were among the first components identified as potential candidates for a subunit vaccine. Subsequent studies have borne out this potential. Monoclonal antibodies (MAbs) directed against rhoptry-associated proteins have been shown to provide substantial inhibition of parasite invasion in vitro (7,17,19,22,29,37). In addition, protein preparations containing both RAP1 and RAP2 have been used to immunize Saimiri monkeys. In these studies, the immunized monkeys developed a lower peak parasitemia than nonimmunized controls and were protected from subsequent lethal blood-stage challenge with P. falciparum (9,30,32,34) in a way similar to the protection acquired by immunization with whole merozoite (27,39,46). Two rhoptry-associated proteins, RAP1 and RAP2, have been sequenced (20,36). Unlike many other P. falciparum antigens, RAP1 and RAP2 exhibit a high degree of sequence similarity among P. falciparum isolates. Isolates from Honduras,

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“…1 Monoclonal antibodies directed against the complex inhibit parasite growth in vitro. [2][3][4][5][6][7] Two vaccine trials in Saimiri sciureus monkeys with RAP1 and RAP2 extracted from parasites using Freund's adjuvants demonstrated that these proteins may induce protective immunity. [8][9][10][11] Monkeys immunized with Plasmodium falciparum parasite-derived RAP1 and RAP2 (PfRAP1 and PfRAP2), isolated by affinity chromatography and further purified by SDS (sodium dodecyl sulfate) polyacrylamide electrophoresis, had substantially lower peak parasitemia when compared to control monkeys and did not require drug treatment.…”

Section: Introductionmentioning

confidence: 99%

Efficacy of vaccines containing rhoptry-associated proteins RAP1 and RAP2 of Plasmodium falciparum in Saimiri boliviensis monkeys.

Collins¹,

Walduck²,

Sullivan³

et al. 2000

The American Journal of Tropical Medicine and Hygiene

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Abstract. A vaccine trial was conducted with rhoptry-associated proteins 1 and 2 (RAP1 and RAP2) of Plasmodium falciparum in Saimiri boliviensis monkeys to compare the ability of parasite-derived (PfRAP1 and 2) and recombinant proteins (rRAP1 and 2) to induce protective immune responses and to find adjuvants suitable for use in humans. Eight groups of 6 monkeys each were immunized with parasite-derived or recombinant RAP1 and 2 with Freund's complete adjuvant (FCA) followed by Freund's incomplete adjuvant (FIA), Montanide ISA720 adjuvant, or CRL1005 adjuvant. Recombinant RAP1 and RAP2 were also administered separately, with Montanide ISA720. After 3 immunizations, monkeys were challenged by iv inoculation of 50,000 parasites of the Uganda Palo Alto strain of P. falciparum. Of the animals vaccinated using FCA/FIA, 1 of 6 control monkeys, 3 of 6 immunized with PfRAP1 and 2, and 2 of 6 with rRAP1 and 2 did not require drug treatment. Of the monkeys vaccinated with Montanide ISA720 adjuvant, 0 of the 6 control monkeys, 2 of 6 immunized with RAP1 and 2, 1 of 6 immunized with rRAP1, and 4 of 6 immunized with RAP2 did not require drug treatment. Two of 6 monkeys immunized with PfRAP1 and 2 with CRL1005 did not require treatment. All groups receiving RAP1, RAP2, or both had a significant decrease in initial parasite multiplication rates and there was a significant negative correlation between anti-RAP2 antibody and multiplication rates. Animals were rechallenged with the homologous parasite 126 days after the first challenge. Of the monkeys that did not require drug treatment after the first challenge, none developed detectable parasitemia following rechallenge.

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Identification and characterisation of proteins associated with the rhoptry organelles of Plasmodium falciparum merozoites

Cited by 55 publications

References 30 publications

Identification and Disruption of the Gene Encoding the Third Member of the Low-Molecular-Mass Rhoptry Complex in Plasmodium falciparum

Identification and Disruption of the Gene Encoding the Third Member of the Low-Molecular-Mass Rhoptry Complex in Plasmodium falciparum

Interaction of HLA and Age on Levels of Antibody toPlasmodium falciparumRhoptry-Associated Proteins 1 and 2

Efficacy of vaccines containing rhoptry-associated proteins RAP1 and RAP2 of Plasmodium falciparum in Saimiri boliviensis monkeys.

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