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PubMed2004

Host cell invasion by the apicomplexans: the significance of microneme protein proteolysis.

Intracellular life-style has been adopted by many pathogens as a successful immune evasion mechanism. To gain entry to a large variety of host cells and to establish an intracellular niche, Toxoplasma gondii and other apicomplexans rely on an active process distinct from phagocytosis. Calcium-regulated secretion of microneme proteins and parasite actin polymerization together with the action of at least one myosin motor act in concert to generate the gliding motility necessary to propel the parasite into host cells. During this active penetration, host cell transmembrane proteins are excluded from the forming parasitophorous vacuole hence conferring the resistance to acidification and degradative fusion. Apicomplexans possess a large repertoire of microneme proteins that contribute to invasion, but their precise role and the level of functional redundancy remain to be evaluated. Remarkably, most microneme proteins are proteolytically cleaved during biogenesis and post-exocytosis. The significance of the processing events and the identification of the proteases implicated are the object of intensive investigations. These proteases may constitute potential drug targets for intervention against malaria and other diseases caused by these parasites.

Animals↗
PubMed2005

Organelle-specific cochaperonins in apicomplexan parasites.

Protein maturation in eukaryotic organelles requires the type I chaperonin system; this comprises chaperonin 60 (Cpn60) and its cochaperonin. We have re-examined and revised the sequence of the nuclear genes specifying organellar cochaperonins in Plasmodium falciparum (Pf). One gene encodes a typical cochaperonin (PfCpn10) whereas the other (encoding PfCpn20) specifies two Cpn10 domains arranged in tandem as in plant chloroplasts. Transfection experiments using fluorescent reporters showed specific localization of PfCpn10 to the mitochondrion and PfCpn20 to the plastid. As P. falciparum also has two Cpn60s, one of which is targeted specifically to the mitochondrion and the other exclusively to the plastid, each organelle has a distinct type I chaperonin system. Comparative sequence analysis extended these findings to several other apicomplexan parasites that have both a mitochondrion and a plastid. Phylogenetic analysis suggests the Cpn10s and Cpn20s of apicomplexans are independently monophyletic. The apicomplexan Cpn10 is phylogenetically related to other mitochondrial versions but a significant relationship between apicomplexan Cpn20s and other cochaperonins was not established.

Amino Acid Sequence↗
PubMed2005

Ultrastructural aspects of Fallisia effusa (Haemosporina: Garniidae) in thrombocytes of the lizard Neusticurus bicarinatus (Reptilia: Teiidae).

The fine structure of the different stages of the Fallisia effusa (Haemosporina: Garniidae), infecting the thrombocytes of the semi-aquatic Amazonian lizard Neusticurus bicarinatus (Reptilia: Teiidae) is described. Gametocytes, meronts, and merozoites of Fallisia effusa were found within a parasitophorous vacuole (PV). Multiple infections of micro- and macrogametocytes were observed. A circumferential coil of microtubules was seen in the cytoplasm of the infected host cell and this microtubule array was pronounced in cells harboring gametocytes. A deep invagination of the inner membrane complex of gametocytes may be involved in nutrition. The non-pigmented parasites underwent both merogony and gametogony in thrombocytes of the peripheral blood. No infection of the erythrocytes was observed. These observations confirm that Fallisia effusa displays characteristic features distinguishing it from other members of the Haemosporidian families, and that it has the ability to modulate microtubule assembly.

Animals↗
PubMed2006

EFL GTPase in cryptomonads and the distribution of EFL and EF-1alpha in chromalveolates.

EFL (EF-like protein) is a member of the GTPase superfamily that includes several translation factors. Because it has only been found in a few eukaryotic lineages and its presence correlates with the absence of the related core translation factor EF-1alpha, its distribution is hypothesized to be the result of lateral gene transfer and replacement of EF-1alpha. In one supergroup of eukaryotes, the chromalveolates, two major lineages were found to contain EFL (dinoflagellates and haptophytes), while the others encode EF-1alpha (apicomplexans, ciliates, heterokonts and cryptomonads). For each of these groups, this distribution was deduced from whole genome sequence or expressed sequence tag (EST) data from several species, with the exception of cryptomonads from which only a single EF-1alpha PCR product from one species was known. By sequencing ESTs from two cryptomonads, Guillardia theta and Rhodomonas salina, and searching for all GTPase translation factors, we revealed that EFL is present in both species, but, contrary to expectations, we found EF-1alpha in neither. On balance, we suggest the previously reported EF-1alpha from Rhodomonas salina is likely an artefact of contamination. We also identified EFL in EST data from two members of the dinoflagellate lineage, Karlodinium micrum and Oxyrrhis marina, and from an ongoing genomic sequence project from a third, Perkinsus marinus. Karlodinium micrum is a symbiotic pairing of two lineages that would have both had EFL (a dinoflagellate and a haptophyte), but only the dinoflagellate gene remains. Oxyrrhis marina and Perkinsus marinus are early diverging sister-groups to dinoflagellates, and together show that EFL originated early in this lineage. Phylogenetic analysis confirmed that these genes are all EFL homologues, and showed that cryptomonad genes are not detectably related to EFL from other chromalveolates, which collectively form several distinct groups. The known distribution of EFL now includes a third group of chromalveolates, cryptomonads. Of the six major subgroups of chromalveolates, EFL is found in half and EF-1alpha in the other half, and none as yet unambiguously possess both genes. Phylogenetic analysis indicates EFL likely arose early within each subgroup where it is found, but suggests it may have originated multiple times within chromalveolates as a whole.

Animals↗
PubMed2004

Molecular and functional aspects of parasite invasion.

Apicomplexan parasites have evolved an efficient mechanism to gain entry into non-phagocytic cells, hence challenging their hosts by the establishment of infection in immuno-privileged tissues. Gliding motility is a prerequisite for the invasive stage of most apicomplexans, allowing them to migrate across tissues, and actively invade and egress host cells. In the late 1960s, detailed morphological studies revealed that motile apicomplexans share an elaborate architecture comprising a subpellicular cytoskeleton and apical organelles. Since 1993, the development of technologies for transient and stable transfection have provided powerful tools with which to identify gene products associated with these structures and organelles, as well as to understand their functions. In combination with access to several parasite genomes, it is now possible to compare and contrast the strategies and molecular machines that have been selectively designed by distinct life stages within a species, or by different apicomplexan species, to optimize infection.

Animals↗
PubMed2006

Why are plastid genomes retained in non-photosynthetic organisms?

The evolution of the plastid from a photosynthetic bacterial endosymbiont involved a dramatic reduction in the complexity of the plastid genome, with many genes either discarded or transferred to the nucleus of the eukaryotic host. However, this evolutionary process has not gone to completion and a subset of genes remains in all plastids examined to date. The various hypotheses put forward to explain the retention of the plastid genome have tended to focus on the need for photosynthetic organisms to retain a genetic system in the chloroplast, and they fail to explain why heterotrophic plants and algae, and the apicomplexan parasites all retain a genome in their non-photosynthetic plastids. Here we consider two additional explanations: the 'essential tRNAs' hypothesis and the 'transfer-window' hypothesis.

Animals↗
PubMed2006

Expressed sequence tag (EST) analysis of the erythrocytic stages of Babesia bovis.

Expressed sequence tags (ESTs) provide an efficient way to identify large numbers of genes expressed in a specific stage of the life cycle of an organism. Here we analysed approximately 13,000 ESTs derived from the erythrocytic stage of the apicomplexan parasite Babesia bovis. The ESTs were clustered in order to obtain information on the expression level of a gene and to increase sequence length and reliability. A total of 3522 clusters were obtained and annotated using BLAST algorithms. The clusters were estimated to represent approximately 2600 genes of which in total approximately 2.1 Mbp sequence information was obtained. Expression levels of the genes, as determined by the numbers of ESTs contained within a cluster, were compared to those of their closest homologs in the erythrocytic stage of Plasmodium falciparum and Toxoplasma gondii tachyzoites. Pathways that are represented relatively abundant in B. bovis are, amongst others, the purine salvage pathway (displaying characteristics not identified before in apicomplexans), isoprenoid biosynthesis in the apicoplast and many genes encoding mitochondrial proteins. Especially remarkable in the latter group are the F-type ATPases - which are hardly expressed in P. falciparum and T. gondii - and two highly expressed glycerol-3-phosphate dehydrogenases creating a shuttle possibly controlling the cytoplasmic NADH/NAD+ -ratio. A comparison of known antigenic proteins from Australian and American strains of B. bovis with the Israel strain used here identifies considerable sequence variation in the rhoptry associated protein-1 (RAP-1), merozoite surface proteins of the variable merozoite surface antigen (VMSA) family and spherical body proteins. Analysis of the EST clusters representing the variable erythocyte surface antigen family reveals many variant transcripts of which a few are dominant. Two putative pseudogenes also seem to be transcribed at high levels.

Animals↗
PubMed1989

Mass spectrometric identification of peptides present in immunized and parasitised hemolymph from honeybees without purification.

We propose that a substance, identified using mass spectrometry, present in the hemolymph of the honeybee (Apis mellifera) as a result of stimulating the insect immune system corresponds with Apidaecin I, an antibacterial peptide recently described. The plasma desorption mass spectra indicate that several other higher molecule mass substances are synthesised as a result of bacterial or parasite infection.

Animals↗
PubMed1985

Theileria parva: expression of a sporozoite surface coat antigen.

A monoclonal antibody specific for the Theileria parva sporozoite, which recognizes a determinant on the surface coat and blocks sporozoite infectivity, was used to investigate the presence of the determinant on other stages of the parasite lifecycle. Immunofluorescence techniques did not demonstrate this determinant on the kinete, schizont, merozoite, or piroplasm stages of the parasite. Immunoautoradiography, using a tritiated form of the monoclonal antibody, on sections of infected salivary glands collected from ticks that had fed for 0, 1, 2, 3, or 4 days revealed that the determinant recognized was synthesized predominantly during sporogony, between 2 to 3 days after the tick started feeding. Immunoelectron microscopy was performed on ultrathin frozen sections of infected tick salivary glands incubated with the monoclonal antibody followed by Protein-A--colloidal gold. The antigen or its precursor could be detected in the developing parasite. In ticks fed 2 days, the sporoblast was labeled, both in the cytoplasm and on parasite membranes, often including the nuclear envelope. In sections from ticks fed 4 days, the sporozoite surface membrane was labeled, as were membrane-bounded sporozoite organelles identified as micronemes. Observation by immunofluorescence, on sporozoites incubated with bovine peripheral blood lymphocytes, suggested that the antigen recognized by the monoclonal antibody does not enter the lymphocyte during sporozoite endocytosis. We conclude that synthesis of the antigen or its precursor(s) occurs during sporogony in the feeding tick, at the time of maximal parasite proliferation, and precedes the formation of morphologically mature sporozoites; the antigen's role in the parasite life cycle also appears to be limited to events associated with the sporozoite entry process.

Animals↗