Biomedical subjects

P R Preiser

Publications and source records attributed to P R Preiser.

16 recordsLinked to original sources

PubMed2004

STEVOR--a multifunctional protein?

The stevor multigene family is the third largest identified in Plasmodium falciparum. Its members have the potential to be involved in antigenic variation and virulence by analogy with the var and rif multigene families. This review highlights recent studies of stevor transcription and expression which show that stevor is distinct from both the var and rif multigene families. STEVOR is expressed during several stages of the lifecycle, and thus may contribute significantly to the long term survival of the parasite.

Animals↗

PubMed2002

Stage-specific transcription of distinct repertoires of a multigene family during Plasmodium life cycle.

Members of a multigene family in the rodent malaria parasite Plasmodium yoelii yoelii code for 235-kilodalton proteins (Py235) that are located in the merozoite apical complex, are implicated in virulence, and may determine red blood cell specificity. We show that distinct subsets of py235 genes are expressed in sporozoites and hepatic and erythrocytic stages. Antibodies to Py235 inhibited sporozoite invasion of hepatocytes. The switch in expression profile occurred immediately after transition from one stage to another. The results suggest that this differential expression is driven by strong biological requirements and provide evidence that hepatic and erythrocytic merozoites differ.

Amino Acid Sequence↗

PubMed2002

The plastid DNA of the malaria parasite Plasmodium falciparum is replicated by two mechanisms.

In common with other apicomplexan parasites, Plasmodium falciparum, a causative organism of human malaria, harbours a residual plastid derived from an ancient secondary endosymbiotic acquisition of an alga. The function of the 35 kb plastid genome is unknown, but its evolutionary origin and genetic content make it a likely target for chemotherapy. Pulsed field gel electrophoresis and ionizing radiation have shown that essentially all the plastid DNA comprises covalently closed circular monomers, together with a tiny minority of linear 35 kb molecules. Using two-dimensional gels and electron microscopy, two replication mechanisms have been revealed. One, sensitive to the topoisomerase inhibitor ciprofloxacin, appears to initiate at twin D-loops located in a large inverted repeat carrying duplicated rRNA and tRNA genes, whereas the second, less drug sensitive, probably involves rolling circles that initiate outside the inverted repeat.

Animals↗

PubMed2002

Small variant STEVOR antigen is uniquely located within Maurer's clefts in Plasmodium falciparum-infected red blood cells.

Malaria parasite antigens encoded by multigene families are important factors in virulence and in disease pathology. In Plasmodium falciparum, the virulence factor PfEMP-1 is encoded by the var multigene family and is exposed at the infected erythrocyte surface. PfEMP-1 is clonally variant, allowing the parasite to evade host immunity. The recently identified P. falciparum stevor multigene family and its products also have the potential to be involved in similar important aspects of host-parasite interactions. Here, we show tightly regulated stage-specific transcription of stevor occurring over just a few hours of the asexual parasite life cycle. Only a subset of stevor genes are transcribed in parasite populations maintained in cultures and in single micromanipulated parasites. Antibodies against STEVOR recognize proteins of the expected size (approximately 37 kDa) and localize STEVOR in Maurer's clefts, unique membranous structures located in the cytoplasm of infected erythrocytes. The fact that the timing of stevor expression and the location of STEVOR are clearly distinct from those of other parasite variant antigens suggests that this gene family may have a novel role in P. falciparum biology.

Animals↗

PubMed2001

The 235 kDa rhoptry protein of Plasmodium (yoelii) yoelii: function at the junction.

All malaria parasites are obligate intracellular organisms that must clearly recognise and discriminate between different cells during their life cycle. Invasion into a cell is a multi-step event that is marked by initial attachment proceeding to irreversible junction formation and penetration. A 235 kDa rhoptry protein (Py235) in the rodent malaria, Plasmodium yoelii yoelii has been shown to be involved in red blood cell (rbc) binding and is involved in a new mechanism of clonal phenotypic variation that may be important in adaptation and immune evasion. Immunisation studies using Py235 have also revealed a role for this protein in the virulence phenotype seen with P. y. yoelii in laboratory mice. Interestingly, the genes that encode this protein are present as a multi-gene family. In this paper, we examine Py235 at the level of DNA, transcription and expression, discussing the role of this protein during invasion, in virulence and in immune evasion.

Animals↗

PubMed2001

An alteration in concatameric structure is associated with efficient segregation of plasmids in transfected Plasmodium falciparum parasites.

Transfection of the human malaria parasite Plasmodium falciparum is currently performed with circularised plasmids that are maintained episomally in parasites under drug selection but which are rapidly lost when selection pressure is removed. In this paper, we show that in instances where gene targeting is not favoured, transfected plasmids can change to stably replicating forms (SRFs) that are maintained episomally in the absence of drug selection. SRF DNA is a large concatamer of the parental plasmid comprising at least nine plasmids arranged in a head-to-tail array. We show as well that the original unstable replicating forms (URFs) are also present as head-to-tail concatamers, but only comprise three plasmids. Limited digestion and gamma irradiation experiments revealed that while URF concatamers are primarily circular, as expected, SRF concatamers form a more complex structure that includes extensive single-stranded DNA. No evidence of sequence rearrangement or additional sequence was detected in SRF DNA, including in transient replication experiments designed to select for more efficiently replicating plasmids. Surprisingly, these experiments revealed that the bacterial plasmid alone can replicate in parasites. Together, these results imply that transfected plasmids are required to form head-to-tail concatamers to be maintained in parasites and implicate both rolling-circle and recombination-dependent mechanisms in their replication.

Animals↗

PubMed2001

Distribution and characterisation of the 235 kDa rhoptry multigene family within the genomes of virulent and avirulent lines of Plasmodium yoelii.

In the rodent malaria species, Plasmodium yoelii, a multi-gene family (Py235) encodes a 235 kDa rhoptry protein. This protein is believed to be involved in merozoite attachment and invasion of red blood cells. Only two members of Py235 have been sequenced so far. Using genomic DNA from the virulent P. yoelii YM line we have PCR amplified additional members of this gene family. These >8 kb full length clones have been cloned and sequenced. Based on differences within the tri-amino acid repeat structure at the C-terminal end of the Py235 protein, it has been possible to divide the multi-gene family into subtypes. The protein translations of five full-length genes (representing four different subtypes) were compared. While there was a high level of amino acid identity at the C-terminal end of these proteins, the N-terminal region revealed a great deal of sequence diversity. Critically, certain residues appeared to be conserved notably seven out of eight cysteines. Comparison of two full-length genes of a particular sub-type shows >99% amino acid identity at the protein level, implying that very closely related genes exist within the parasite genome. We have used this new sequence information to compare the distribution of Py235 in the virulent YM and avirulent 17X lines of P. yoelii. Our results indicate that while the overall distribution of Py235 genes is broadly conserved between the two lines, significant differences exist when individual subtypes are compared.

Amino Acid Sequence↗

PubMed2000

Malaria multigene families: the price of chronicity.

In this article, Georges Snounou, William Jarra and Peter Preiser discuss the survival strategy of malaria parasites in the light of a novel mechanism of clonal phenotypic variation recently described for a multigene family of Plasmodium yoelii yoelii. The 235 kDa rhoptry proteins (Py235) encoded by these genes may be involved in the selection of red blood cells for invasion by merozoites. The new mechanism may explain the ability of individual parasites to adapt to natural variations in red blood cell subsets, while ensuring that sufficient merozoites escape immune attack, thus maintaining a chronic infection for extended periods. This counterpoints the antigenic variation exemplified by PfEMP1 proteins (a large family of proteins derived from P. falciparum), which operates at the population level. The possibility of manipulating the expression of functionally similar genes in other Plasmodium species could lead to therapies aimed at reducing clinical severity without compromising the acquisition and maintenance of immunity.

Animals↗

PubMed1999

A rhoptry-protein-associated mechanism of clonal phenotypic variation in rodent malaria.

The recognition and invasion of host cells are mediated by components of the apical complex of the ookinete, sporozoite and merozoite stages of Plasmodium parasites. The paired rhoptries (organelles involved in host-cell recognition) in the apical complex contain many proteins of as-yet unknown function. In the rodent malaria agent P. yoelii yoelii, a multigene family codes for merozoite rhoptry proteins of relative molecular mass 235,000 (p235 proteins); these proteins are thought to determine the subset of erythrocytes that the parasites invade. Further support for this idea came from the identification of a region in p235 with weak but significant homology to reticulocyte-binding protein-2 of P. vivax and the demonstration that at least one p235 member binds to the erythrocyte surface membrane. Here, using single, micromanipulated P.y.yoelii parasites, we describe a new mechanism of gene expression by which the merozoites originating from a single schizont each express a distinct member of this multigene family. We propose that this new type of clonal phenotypic variation provides the parasite with a survival strategy in the mammalian host; this strategy contributes to the observed chronicity of malarial infections. This phenomenon is genetically and functionally distinct from classical antigenic variation, which is mediated by the var multigene family of P. falciparum.

Animals↗

PubMed1999

Antibiotic inhibitors of organellar protein synthesis in Plasmodium falciparum.

Elongation factor Tu (EF-Tu) is encoded by the tuf gene of the plastid organelle of the malaria parasite Plasmodium falciparum. A range of structurally unrelated inhibitors of this GTP-dependent translation factor was shown to have antimalarial activity in blood cultures. The most active was the cyclic thiazolyl peptide amythiamicin A with an IC50 = 0.01 microM. Demonstrable complexes were formed in vitro between a recombinant version of P. falciparum EF-Tu(pl) and inhibitors that bind to different sites on EF-Tu; these included the antibiotics kirromycin, GE2270A and enacyloxin IIa.

Animals↗

PubMed1998

Plasmodium yoelii: differences in the transcription of the 235-kDa rhoptry protein multigene family in lethal and nonlethal lines.

We have compared the transcription of the 235-kDa rhoptry protein (p235) multigene family in the lethal (YM) and nonlethal (17X) lines of the rodent malaria parasite Plasmodium yoelii. This protein is thought to be involved in erythrocyte invasion by the parasite. Using a PCR-based approach we demonstrated that both lines have similar p235 families. However, RT-PCR analysis revealed that this similarity is not evident at the level of transcription, with the lethal line not transcribing a whole subset of its p235 gene repetoire. Specific anti-p235 immune pressure induces differences in invasion properties of the lethal line; we were, however, unable to detect any changes in the transcription pattern of the p235 genes associated with this event.

Amino Acid Sequence↗

PubMed1996

Complete gene map of the plastid-like DNA of the malaria parasite Plasmodium falciparum.

Malaria parasites, and other parasitic protists of the Phylum Apicomplexa, carry a plastid-like genome with greatly reduced sequence complexity. This 35 kb DNA circle resembles the plastid DNA of non-photosynthetic plants, encoding almost exclusively components involved in gene expression. The complete gene map described here includes genes for duplicated large and small subunit rRNAs, 25 species of tRNA, three subunits of a eubacterial RNA polymerase, 17 ribosomal proteins, and a translation elongation factor. In addition, it codes for an unusual member of the Clp family of chaperones, as well as an open reading frame of unknown function found in red algal plastids. Transcription is polycistronic. This plastid-like DNA molecule is conserved in several genera of apicomplexans and is conjectured to have been acquired by an early progenitor of the Phylum by secondary endosymbiosis. The function of the organelle (plastid) carrying this DNA remains obscure, but appears to be specified by genes transferred to the nucleus.

Amino Acid Sequence↗

PubMed1996

Recombination associated with replication of malarial mitochondrial DNA.

Mitochondrial DNA of the malarial parasite Plasmodium falciparum comprises approximately 20 copies per cell of a 6 kb genome, arranged mainly as polydisperse linear concatemers. In synchronous blood cultures, initiation of mtDNA replication coincides with the start of the 4-5 doublings in nuclear DNA that mark the reproductive phase of the erythrocytic cycle. We show that mtDNA replication coincides with a recombination process reminiscent of the replication mechanism used by certain bacteriophages and plasmids. The few circular forms of mtDNA which are also present do not replicate by a theta mechanism, but are themselves the product of recombination, and we propose they undergo rolling circle activity to generate the linear concatemers.

Animals↗

PubMed1996

Organelle DNAs: The bit players in malaria parasite DNA replication.

The replication mechanics of the extrachromosomal DNAs of the malaria parasite are beginning to be anravelled. At 6 kb, the mitochondrial genome is the smallest known and, unlike higher eukaryotes, its multiple copies per cell occur as polydisperse linear concatemers. Here, Don Williamson, Peter Preiser and Iain Wilson discuss recent evidence that this DNA replicates by a process akin to those of certain bacteriophages, which make use of extensive recombination coupled with rolling circles. The parasite's second extrachromosomal DNA, a 35 kb circular molecule thought to be a plastid remnant inherited from a remote photoautotroph, probably replicates in a more familiar fashion from conventional origins or D loops. Improved understanding of both organelle's replicative mechanisms could give new leads to malaria chemotherapy.

Journal Article↗

PubMed1991

Drosophila 5 S RNA processing requires the 1-118 base pair and additional sequence proximal to the processing site.

Using an in vitro processing system, we have identified a required sequence surrounding the Drosophila melanogaster 5 S RNA processing site at nucleotide 120. Mutations in this region vary the processing rate from complete inhibition to a level equal to or greater than wild type. Analysis of mutants at +1 and in the region 118-122 separates the inhibitory effect into two parts. 1) Nucleotide 118 C, the base-paired nucleotide in helix I proximal to the processing site, plays an essential role. Changing it to a purine inhibits processing. The +1-118 base pair must be intact, but this alone is not sufficient for processing, since compensatory changes at +1 do not restore down-processing mutants at 118 to the wild type level. 2) The processing site has to be pyrimidine rich; multiple contiguous purines inhibit processing. On the other hand, multiple pyrimidines can largely negate the inhibitory effect of a mutation at position 118. Thus a base-paired C at 118 followed by a stretch of pyrimidines is the processing signal, which may be recognized by the processing enzyme and/or a required accessory factor.

Animals↗

PubMed1991

In vitro processing of Drosophila melanogaster 5 S ribosomal RNA. 3' end effects and requirement for internal domains of mature 5 S RNA.

5 S RNA processing in Drosophila melanogaster, the removal of 15 nucleotides from the 3' end of the 135-nucleotide (nt) primary transcript, may play an important role in the regulation of 5 S RNA transport and ribosome assembly. We have uncoupled processing from transcription using gel purified primary transcripts processed in vitro by a cellular S100 extract. The RNA was generated by a homologous transcription system or by a T7 RNA polymerase reaction using a constructed 5 S RNA gene linked to a T7 promoter. In vitro D. melanogaster 5 S RNA processing is heat- and EDTA-sensitive, suggesting a requirement for protein, and produces a 3' end characteristic of mature 5 S RNA. Processing of substrate RNAs with altered 3' ends shows that the 3'-U5 tail (nt 131-135) inhibits the reaction. 30 nt, including all of domain IV and most of domain V, are dispensible for processing, whereas deletions including the base of stem V and all or part of stem III severely inhibit it. Several possible mechanisms are discussed.

Animals↗