RESUMO
Most studies of gene expression in Plasmodium have been concerned with asexual and/or sexual erythrocytic stages. Identification and cloning of genes expressed in the preerythrocytic stages lag far behind. We have constructed a high quality cDNA library of the Plasmodium sporozoite stage by using the rodent malaria parasite P. yoelii, an important model for malaria vaccine development. The technical obstacles associated with limited amounts of RNA material were overcome by PCR-amplifying the transcriptome before cloning. Contamination with mosquito RNA was negligible. Generation of 1,972 expressed sequence tags (EST) resulted in a total of 1,547 unique sequences, allowing insight into sporozoite gene expression. The circumsporozoite protein (CS) and the sporozoite surface protein 2 (SSP2) are well represented in the data set. A BLASTX search with all tags of the nonredundant protein database gave only 161 unique significant matches (P(N) < or = 10(-4)), whereas 1,386 of the unique sequences represented novel sporozoite-expressed genes. We identified ESTs for three proteins that may be involved in host cell invasion and documented their expression in sporozoites. These data should facilitate our understanding of the preerythrocytic Plasmodium life cycle stages and the development of preerythrocytic vaccines.
Assuntos
Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Biblioteca Gênica , Plasmodium yoelii/genética , RNA Mensageiro/biossíntese , RNA de Protozoário/biossíntese , Transcrição Gênica , Motivos de Aminoácidos , Sequência de Aminoácidos , Animais , Anopheles/parasitologia , DNA Complementar/genética , Etiquetas de Sequências Expressas , Interações Hospedeiro-Parasita/genética , Ligantes , Vacinas Antimaláricas , Dados de Sequência Molecular , Plasmodium falciparum/genética , Plasmodium yoelii/crescimento & desenvolvimento , Plasmodium yoelii/patogenicidade , Proteínas de Protozoários/genética , RNA Mensageiro/genética , RNA de Protozoário/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Especificidade da Espécie , Virulência/genéticaRESUMO
It is anticipated that the sequencing of Plasmodium falciparum genome will soon be completed. Rodent models of malaria infection and stable transformation systems provide powerful means of using this information to study gene function in vivo. To date, gene targeting has only been developed for one rodent malaria species, Plasmodium berghei. Another rodent species, Plasmodium yoelii, however, is favored to study the mechanisms of protective immunity to the pre-erythrocytic stages of infection and vaccine development. In addition, it offers the opportunity to investigate unique aspects of pathogenesis of blood stage infection. Here, we report on the stable transfection and gene targeting of P. yoelii. Purified late blood stage schizonts were used as targets for electroporation with a plasmid that contains a pyrimethamine-resistant form of the P. berghei dihydrofolate reductase-thymidylate synthase (Pbdhfr-ts) fused to green fluorescent protein (gfp) gene. After drug selection, fluorescent parasites contained intact, non-rearranged plasmids that remain stable under drug-pressure. In addition, we used another dhfr-ts/gfp based plasmid to disrupt the P. yoelii trap (thrombospondin-related anonymous protein) locus by site-specific integration. The phenotype of P. yoelii TRAP knockout was identical to that previously reported for the P. berghei TRAP knockout. In the absence of TRAP, the erythrocytic cycle, gametocyte and oocyst development of the mutant parasites were indistinguishable from wild type (WT). Although the sporozoites appeared morphologically normal, they failed to glide and to invade the salivary glands of mosquitoes.
Assuntos
Malária/parasitologia , Plasmodium yoelii/genética , Transfecção , Transformação Genética , Animais , Antimaláricos/farmacologia , Deleção de Genes , Marcação de Genes , Camundongos , Camundongos Endogâmicos BALB C , Complexos Multienzimáticos/efeitos dos fármacos , Complexos Multienzimáticos/genética , Fenótipo , Plasmídeos/genética , Plasmodium yoelii/efeitos dos fármacos , Plasmodium yoelii/crescimento & desenvolvimento , Plasmodium yoelii/patogenicidade , Proteínas de Protozoários/genética , Pirimetamina/farmacologia , Tetra-Hidrofolato Desidrogenase/efeitos dos fármacos , Tetra-Hidrofolato Desidrogenase/genética , Timidilato Sintase/efeitos dos fármacos , Timidilato Sintase/genéticaRESUMO
Trypanosoma cruzi trypomastigotes treated with phosphatidylinositol-specific phospholipase C (PI-PLC) in vitro are rapidly induced to differentiate into round forms. Using confocal microscopy, we were able to show that trypomastigotes treated with PI-PLC initiate the process of flagellum remodeling by 30 sec after contact with the enzyme and amastigote-like forms are detected as early as 10 min after PI-PLC treatment. Scanning and transmission electron microscopy indicate that trypomastigotes undergo a previously undescribed process of flagellum circularization and internalization. Analysis of the flagellar complex with monoclonal antibody 4D9 shows heterogeneous labeling among the parasites, suggesting a remodeling of these molecules. After PI-PLC treatment, parasites rapidly lose the surface marker Ssp-3 and 24 h post-treatment they begin to exhibit a circular nucleus and a rod-shaped kinetoplast. By flow cytometry analysis and confocal microscopy, the Ssp-4 amastigote-specific epitope can be detected on the parasite surface. This indicates that the release of trypomastigote GPI-anchored molecules by exogenous PI-PLC in vitro can trigger morphological changes.
Assuntos
Glicosilfosfatidilinositóis/metabolismo , Trypanosoma cruzi/citologia , Trypanosoma cruzi/metabolismo , Fosfolipases Tipo C/metabolismo , Animais , Citoesqueleto/ultraestrutura , Flagelos/fisiologia , Microscopia Confocal , Microscopia Eletrônica de Varredura , Fosfatidilinositol Diacilglicerol-Liase , Fosfoinositídeo Fosfolipase C , Trypanosoma cruzi/crescimento & desenvolvimentoRESUMO
Previously we have used the Plasmodium dihydrofolate reductase thymidylate synthase (DHFR-TS) selectable marker to generate Plasmodium berghei TRAP null mutant parasites. These TRAP null mutants do not glide and they showed a great reduction in their ability to infect mosquito salivary glands and the hepatocytes of the vertebrate host. Thus far, complementation of these knockout parasites was not possible due to the lack of additional selectable markers. Recently, a new selectable marker, based on the human dihydrofolate reductase (hDHFR) gene, has been developed which confers resistance to the antifolate drug WR99210. This drug has been found to be highly active against pyrimethamine-sensitive and -resistant strains of P. berghei. In this study, we have used the hDHFR gene as a second selectable marker for the complementation of P. berghei TRAP null mutant parasites. Restoration of the TRAP null mutant parasites to the wild-type phenotype was achieved in this study via autonomously replicating episomes bearing a wild-type copy of the TRAP gene. This is the first report of complementation of a mutant phenotype in malaria parasites.
Assuntos
Plasmodium berghei/genética , Proteínas de Protozoários/genética , Tetra-Hidrofolato Desidrogenase/genética , Animais , Antagonistas do Ácido Fólico/farmacologia , Marcadores Genéticos , Humanos , Mutação , Plasmodium berghei/efeitos dos fármacos , Proteínas de Protozoários/metabolismo , Sensibilidade e Especificidade , TransfecçãoRESUMO
Here, we document for the first time the presence of the 26S proteasome and the ubiquitin pathway in a protozoan parasite that is in an early branch in the eukaryotic lineage. The 26S proteasome of Trypanosoma cruzi epimastigotes was identified as a high molecular weight complex (1400 kDa) with an ATP-dependent chymotrypsin-like activity against the substrate Suc-LLVY-Amc. This activity was inhibited by proteasome inhibitors and showed same electrophorectic migration pattern as yeast 26S proteasome in nondenaturating gels. About 30 proteins in a range of 25-110 kDa were detected in the purified T. cruzi 26S proteasome. Antibodies raised against the AAA family of ATPases from eukaryotic 26S proteasome and the T. cruzi 20S core specifically recognized components of T. cruzi 26S. To confirm the biological role of 26S in this primitive eukaryotic parasite, we analyzed the participation of the ubiquitin (Ub)-proteasome system in protein degradation during the time of parasite remodeling. Protein turnover in trypomastigotes was proteasome and ATP-dependent and was enhanced during the transformation of the parasites into amastigotes. If 20S proteasome activity is inhibited, ubiquitinated proteins accumulate in the parasites. As expected from the profound morphological changes that occur during transformation, cytoskeletal proteins associated with the flagellum are targets of the ubiquitin-proteasome pathway.
Assuntos
Acetilcisteína/análogos & derivados , Peptídeo Hidrolases/fisiologia , Proteínas de Protozoários/fisiologia , Trypanosoma cruzi/crescimento & desenvolvimento , Trypanosoma cruzi/metabolismo , Ubiquitinas/fisiologia , Acetilcisteína/farmacologia , Trifosfato de Adenosina/fisiologia , Animais , Linhagem Celular , Permeabilidade da Membrana Celular , Cisteína Endopeptidases/isolamento & purificação , Cisteína Endopeptidases/metabolismo , Cisteína Endopeptidases/fisiologia , Inibidores de Cisteína Proteinase/farmacologia , Proteínas do Citoesqueleto/metabolismo , Flagelos/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Hidrólise , Cinética , Macaca mulatta , Complexos Multienzimáticos/isolamento & purificação , Complexos Multienzimáticos/metabolismo , Complexos Multienzimáticos/fisiologia , Peptídeo Hidrolases/isolamento & purificação , Peptídeo Hidrolases/metabolismo , Complexo de Endopeptidases do Proteassoma , Proteínas de Protozoários/isolamento & purificação , Proteínas de Protozoários/metabolismo , Trypanosoma cruzi/enzimologia , Ubiquitinas/metabolismoAssuntos
Malária/parasitologia , Miosinas/genética , Plasmodium berghei/fisiologia , Plasmodium yoelii/fisiologia , Proteínas de Protozoários/genética , Sequência de Aminoácidos , Animais , Hepatócitos/parasitologia , Dados de Sequência Molecular , Miosinas/química , Miosinas/metabolismo , Plasmodium berghei/patogenicidade , Plasmodium yoelii/patogenicidade , Proteínas de Protozoários/química , Proteínas de Protozoários/metabolismo , Alinhamento de SequênciaRESUMO
Intracellular bacteria and parasites typically invade host cells through the formation of an internalization vacuole around the invading pathogen. Plasmodium sporozoites, the infective stage of the malaria parasite transmitted by mosquitoes, have an alternative mechanism to enter cells. We observed breaching of the plasma membrane of the host cell followed by rapid repair. This mode of entry did not result in the formation of a vacuole around the sporozoite, and was followed by exit of the parasite from the host cell. Sporozoites traversed the cytosol of several cells before invading a hepatocyte by formation of a parasitophorous vacuole, in which they developed into the next infective stage. Sporozoite migration through several cells in the mammalian host appears to be essential for the completion of the life cycle.
Assuntos
Fluoresceína-5-Isotiocianato/análogos & derivados , Hepatócitos/parasitologia , Plasmodium yoelii/fisiologia , Animais , Linhagem Celular , Membrana Celular/parasitologia , Membrana Celular/fisiologia , Membrana Celular/ultraestrutura , Movimento Celular , Citosol/metabolismo , Citosol/parasitologia , Dextranos/metabolismo , Endocitose , Citometria de Fluxo , Fluoresceína-5-Isotiocianato/metabolismo , Hepatócitos/ultraestrutura , Malária/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Plasmodium/fisiologia , Plasmodium yoelii/crescimento & desenvolvimento , Propídio/metabolismo , Toxoplasma/fisiologia , Células Tumorais Cultivadas , Vacúolos/parasitologia , Vacúolos/ultraestruturaAssuntos
Marcação de Genes , Plasmodium/metabolismo , Proteínas de Protozoários , Animais , Malária/parasitologia , Plasmodium/genética , Plasmodium/crescimento & desenvolvimento , Plasmodium/patogenicidade , Proteínas de Protozoários/química , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Relação Estrutura-AtividadeRESUMO
Thrombospondin-related anonymous protein (TRAP), a candidate malaria vaccine antigen, is required for Plasmodium sporozoite gliding motility and cell invasion. For the first time, the ability of antibodies against TRAP to inhibit sporozoite infectivity in vivo is evaluated in detail. TRAP contains an A-domain, a well-characterized adhesive motif found in integrins. We modeled here a three-dimensional structure of the TRAP A-domain of Plasmodium yoelii and located regions surrounding the MIDAS (metal ion-dependent adhesion site), the presumed business end of the domain. Mice were immunized with constructs containing these A-domain regions but were not protected from sporozoite challenge. Furthermore, monoclonal and rabbit polyclonal antibodies against the A-domain, the conserved N terminus, and the repeat region of TRAP had no effect on the gliding motility or sporozoite infectivity to mice. TRAP is located in micronemes, secretory organelles of apicomplexan parasites. Accordingly, the antibodies tested here stained cytoplasmic TRAP brightly by immunofluorescence. However, very little TRAP could be detected on the surface of sporozoites. In contrast, a dramatic relocalization of TRAP onto the parasite surface occurred when sporozoites were treated with calcium ionophore. This likely mimics the release of TRAP from micronemes when a sporozoite contacts its target cell in vivo. Contact with hepatoma cells in culture also appeared to induce the release of TRAP onto the surface of sporozoites. If large amounts of TRAP are released in close proximity to its cellular receptor(s), effective competitive inhibition by antibodies may be difficult to achieve.
Assuntos
Anticorpos Antiprotozoários/farmacologia , Vacinas Antimaláricas/uso terapêutico , Malária/prevenção & controle , Plasmodium yoelii/imunologia , Proteínas de Protozoários/uso terapêutico , Sequência de Aminoácidos , Animais , Epitopos , Proteínas de Membrana/isolamento & purificação , Modelos Moleculares , Dados de Sequência Molecular , Testes de Neutralização , Fragmentos de Peptídeos/imunologia , Estrutura Terciária de Proteína , Proteínas de Protozoários/genética , Proteínas de Protozoários/imunologia , Proteínas de Protozoários/isolamento & purificação , Vacinação , Virulência/efeitos dos fármacosAssuntos
Plasmodium yoelii/patogenicidade , Receptores Imunológicos/genética , Receptores Imunológicos/fisiologia , Proteínas Virais , Animais , Células Cultivadas , Humanos , Integrases/genética , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade , Malária/etiologia , Malária/genética , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Plasmodium yoelii/crescimento & desenvolvimento , VirulênciaRESUMO
Most Apicomplexan parasites, including the human pathogens Plasmodium, Toxoplasma, and Cryptosporidium, actively invade host cells and display gliding motility, both actions powered by parasite microfilaments. In Plasmodium sporozoites, thrombospondin-related anonymous protein (TRAP), a member of a group of Apicomplexan transmembrane proteins that have common adhesion domains, is necessary for gliding motility and infection of the vertebrate host. Here, we provide genetic evidence that TRAP is directly involved in a capping process that drives both sporozoite gliding and cell invasion. We also demonstrate that TRAP-related proteins in other Apicomplexa fulfill the same function and that their cytoplasmic tails interact with homologous partners in the respective parasite. Therefore, a mechanism of surface redistribution of TRAP-related proteins driving gliding locomotion and cell invasion is conserved among Apicomplexan parasites.
Assuntos
Apicomplexa/fisiologia , Apicomplexa/patogenicidade , Infecções por Protozoários/parasitologia , Antígeno 12E7 , Sequência de Aminoácidos , Substituição de Aminoácidos/genética , Animais , Antígenos CD/química , Antígenos CD/fisiologia , Moléculas de Adesão Celular/química , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/fisiologia , Citoplasma/genética , Citoplasma/fisiologia , Humanos , Dados de Sequência Molecular , Movimento , Peptídeos/metabolismo , Plasmodium berghei/patogenicidade , Plasmodium berghei/fisiologia , Infecções por Protozoários/metabolismo , Proteínas de Protozoários/química , Proteínas de Protozoários/genética , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Deleção de Sequência , Homologia de Sequência de Aminoácidos , Toxoplasma/patogenicidade , Toxoplasma/fisiologiaRESUMO
We present a new marker that confers both resistance to pyrimethamine and green fluorescent protein-based fluorescence on the malarial parasite Plasmodium berghei. A single copy of the cassette integrated into the genome is sufficient to direct fluorescence in parasites throughout the life cycle, in both its mosquito and vertebrate hosts. Erythrocyte stages of the parasite that express the marker can be sorted from control parasites by flow cytometry. Pyrimethamine pressure is not necessary for maintaining the cassette in transformed parasites during their sporogonic cycle in mosquitoes, including when it is borne by a plasmid. This tool should thus prove useful in molecular studies of P. berghei, both for generating parasite variants and monitoring their behavior.
Assuntos
Proteínas Luminescentes/genética , Plasmodium berghei/genética , Transformação Genética , Animais , Antimaláricos/farmacologia , Sequência de Bases , Linhagem Celular , Culicidae/parasitologia , Primers do DNA/genética , Resistência a Medicamentos/genética , Eritrócitos/parasitologia , Fluorescência , Marcadores Genéticos , Proteínas de Fluorescência Verde , Plasmodium berghei/efeitos dos fármacos , Plasmodium berghei/crescimento & desenvolvimento , Pirimetamina/farmacologia , RatosRESUMO
Natural resistance of humans to the cattle pathogen Trypanosoma brucei brucei has been attributed to the presence in human serum of nonimmune factors that lyse the parasite. Normal human serum contains two trypanosome lytic factors (TLFs). TLF1 is a 500-kDa lipoprotein, which is reported to contain apolipoprotein A-I (apoA-I), haptoglobin-related protein (Hpr), hemoglobin, paraoxonase, and apoA-II, whereas TLF2 is a larger, poorly characterized particle. We report here a new immunoaffinity-based purification procedure for TLF2 and TLF1, as well as further characterization of the components of each purified TLF. Immunoaffinity-purified TLF1 has a specific activity 10-fold higher than that of TLF1 purified by previously described methods. Moreover, we find that TLF1 is a lipoprotein particle that contains mainly apoA-I and Hpr, trace amounts of paraoxonase, apoA-II, and haptoglobin, but no detectable hemoglobin. Characterization of TLF2 reveals that it is a 1,000-kDa protein complex containing mainly immunoglobulin M, apoA-I, and Hpr but less than 1% detectable lipid.
Assuntos
Lipoproteínas HDL/imunologia , Trypanosoma brucei brucei/imunologia , Sequência de Aminoácidos , Animais , Western Blotting , Fracionamento Químico , Eletroforese em Gel de Poliacrilamida , Humanos , Imunoglobulina M/imunologia , Lipoproteínas HDL/química , Lipoproteínas HDL/classificação , Lipoproteínas HDL/isolamento & purificação , Dados de Sequência Molecular , Testes de Precipitina , Análise de SequênciaRESUMO
In order to provide a rational basis for the development of a pre-erythrocytic malaria vaccine we have aimed at: (a) elucidating the mechanisms of protection, and (b) identifying vaccine formulations that best elicit protection in experimental animals and humans. Based on earlier successful immunization of experimental animals with irradiated sporozoites, human volunteers were exposed to the bites of large numbers of Plasmodium falciparum or P. vivax infected irradiated mosquitoes. The result of this vaccine trial demonstrated for the first time that a pre-erythrocytic vaccine, administered to humans, can result in their complete resistance to malaria infection. However, since infected irradiated mosquitoes are unavailable for large scale vaccination, the alternative is to develop subunit vaccines. The human trials using irradiated sporozoites provided valuable information on the human immune responses to pre-erythrocytic stages and studies on mice an excellent experimental model to characterize protective immune mechanisms. The circumsporozoite protein, the first pre-erythrocytic antigen identified, is present in all malaria species, displaying a similar structure, with a central region of repeats, and two conserved regions, essential for parasite development. Most pre-erythrocytic vaccine candidates are based on the CS protein, expressed in various cell lines, microorganisms, and recently the corresponding DNA. We and others have identified CS-specific B and T cell epitopes, recognized by the rodent and human immune systems, and used them for the development of synthetic vaccines. We used synthetic peptide vaccines, multiple antigen peptides and polyoximes, for immunization, first in experimental animals, and recently in two human safety and immunogenicity trials. We also report here on our work on T cell mediated immunity, particularly the protection of mice immunized with viral vectors expressing CS-specific cytotoxic CD8+ T cell epitopes, and the striking booster effect of recombinant vaccinia virus. To what degree CD8+ T cells, and/or other T cells specific for sporozoites and/or liver stage epitopes, contribute to pre-erythrocytic protective immunity in humans, remains to be determined.
Assuntos
Vacinas Antimaláricas , Plasmodium falciparum/imunologia , Plasmodium vivax/imunologia , Animais , Anticorpos Antiprotozoários/biossíntese , Linfócitos T CD8-Positivos/imunologia , Ensaios Clínicos como Assunto , Humanos , Imunidade Celular , Interferon gama/metabolismo , CamundongosRESUMO
Proteasomes degrade most of the proteins inside eukaryotic cells, including transcription factors and regulators of cell cycle progression. Here we show that nanomolar concentrations of lactacystin, a specific irreversible inhibitor of the 20S proteasome, inhibit development of the exoerythrocytic and erythrocytic stages of the malaria parasite. Although lactacystin-treated Plasmodium berghei sporozoites are still invasive, their development into exoerythrocytic forms (EEF) is inhibited in vitro and in vivo. Erythrocytic schizogony of P. falciparum in vitro is also profoundly inhibited when drug treatment of the synchronized parasites is prior, but not subsequent, to the initiation of DNA synthesis, suggesting that the inhibitory effect of lactacystin is cell cycle specific. Lactacystin reduces P. berghei parasitemia in rats, but the therapeutic index is very low. Along with other studies showing that lactacystin inhibits stage-specific transformation in Trypanosoma and Entamoeba spp., these findings highlight the potential of proteasome inhibitors as drugs for the treatment of diseases caused by protozoan parasites.
Assuntos
Antimaláricos/farmacologia , Cisteína Endopeptidases/efeitos dos fármacos , Inibidores de Cisteína Proteinase/farmacologia , Complexos Multienzimáticos/efeitos dos fármacos , Plasmodium/efeitos dos fármacos , Acetilcisteína/análogos & derivados , Acetilcisteína/farmacologia , Animais , Eritrócitos/parasitologia , Humanos , Hipoxantina/metabolismo , Plasmodium/crescimento & desenvolvimento , Complexo de Endopeptidases do Proteassoma , Ratos , Ratos Sprague-DawleyRESUMO
Haptoglobin-related protein (HPR) is a serum protein that is >90% homologous to the acute-phase reactant haptoglobin (Hp). Haptoglobin binds and removes free hemoglobin (Hb) from the circulation. Hpr levels are elevated with tumor progression in the serum of some cancer patients, but the relevance of this observation is not understood. HPR is an integral part of two distinct high molecular weight complexes (trypanosome lytic factor 1 (TLF1) and TLF2) that are lytic for the African parasite Trypanosoma brucei brucei. Previous data indicate that HPR represents the toxic component of both trypanosome lytic factors. It has been proposed that after uptake by the parasite, Hb bound to HPR causes lysis in a peroxidase-dependent process. We report that the molecular architecture of HPR in normal human serum is different from that of Hp and that HPR does not bind Hb in normal human serum. Immunodepletion of all detectable Hb from TLF1 does not deplete TLF1 of HPR or trypanolytic activity, suggesting that the mechanism of parasite lysis is Hb-independent.
Assuntos
Antígenos de Neoplasias , Proteínas Sanguíneas/fisiologia , Trypanosoma brucei brucei/metabolismo , Animais , Biomarcadores , Proteínas Sanguíneas/química , Proteínas Sanguíneas/farmacologia , Haptoglobinas/metabolismo , Hemoglobinas/metabolismo , Testes de Precipitina , Trypanosoma brucei brucei/químicaRESUMO
Malaria infection of the host cells requires host-parasite recognition events mediated by adhesion and signaling molecules. Recent development of systems for stable transformation and targeted integration of exogenous DNA in malaria parasites provides a powerful tool to study the structure and function of Plasmodium attachment motifs, and their role in infection and disease.
Assuntos
Moléculas de Adesão Celular/metabolismo , Malária/parasitologia , Plasmodium/patogenicidade , Proteínas de Protozoários/metabolismo , Sequência de Aminoácidos , Animais , Adesão Celular , Moléculas de Adesão Celular/genética , Eritrócitos/parasitologia , Fígado/parasitologia , Dados de Sequência Molecular , Plasmodium/genética , Proteínas de Protozoários/genéticaAssuntos
Antígenos CD/fisiologia , Moléculas de Adesão Celular/fisiologia , Complemento C3/fisiologia , Complemento C4/fisiologia , Molécula 1 de Adesão Intercelular/fisiologia , Malária/etiologia , Plasmodium yoelii/patogenicidade , Sequência de Aminoácidos , Animais , Antígenos CD/genética , Sequência de Bases , Adesão Celular/genética , Adesão Celular/fisiologia , Moléculas de Adesão Celular/genética , Complemento C3/genética , Complemento C4/genética , Primers do DNA/genética , Marcação de Genes , Molécula 1 de Adesão Intercelular/genética , Fígado/parasitologia , Malária/genética , Malária/imunologia , Camundongos , Camundongos Knockout , Plasmodium yoelii/genética , Plasmodium yoelii/fisiologia , Reação em Cadeia da Polimerase , Proteínas de Protozoários/genética , Proteínas de Protozoários/fisiologiaRESUMO
Many protozoans of the phylum Apicomplexa are invasive parasites that exhibit a substrate-dependent gliding motility. Plasmodium (malaria) sporozoites, the stage of the parasite that invades the salivary glands of the mosquito vector and the liver of the vertebrate host, express a surface protein called thrombospondin-related anonymous protein (TRAP) that has homologs in other Apicomplexa. By gene targeting in a rodent Plasmodium, we demonstrate that TRAP is critical for sporozoite infection of the mosquito salivary glands and the rat liver, and is essential for sporozoite gliding motility in vitro. This suggests that in Plasmodium sporozoites, and likely in other Apicomplexa, gliding locomotion and cell invasion have a common molecular basis.