Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0024530 (malaria)
 44,886 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Autoradiography was used to study the incorporation of tritium-labelled nucleic acid precursors into the sporogonic stages of Plasmodium cynomolgi in Anopheles b. balabacensis. Infected mosquitoes were fed on either 3H-adenine (5, 25 and 50 muCi/ml or 3H-thymidine (50 muCi/ml) for various periods after the blood meal. 3H-adenine was incorporated into DNA and RNA of the developing oocysts and the resulting sporozoites. Midgut epithelial cells incorporated label from 3H-adenine into nuclear and cytoplasmic regions. In both parasite and host tissue RNA-label was removed after RNase treatment. 3H-thymidine was not taken up by the developing oocysts or sporozoites while incorporation into the cell nuclei of the adjacent midgut epithelium and fat-body of the mosquito was shown. The observed incorporation of 3H-adenine, but not 3H-thymidine, into the sporogonic stages of Plasmodium supports the assumption that the malaria parasite needs exogenous sources of purine but relies on the de novo synthesis of pyrimidine during its development in the mosquito.
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PMID:Incorporation of nucleic acid precursors by Plasmodium cynomolgi in Anopheles balabacensis. 118 24

CD36 is an 88-kD integral membrane protein expressed on platelets, monocytes, macrophages, certain microvascular endothelia, and retinal pigment epithelium. It functions as an adhesive receptor for thrombospondin-1 (TSP-1), collagen, and malaria-infected erythrocytes and as a scavenger receptor for oxidized LDL and photoreceptor outer segments. The CD36-TSP-1 interaction plays a role in cell adhesion and the phagocytosis of apoptotic cells by macrophages. Because of the potential importance of the CD36-TSP-1 interaction in mediating atherogenic and inflammatory processes, we studied their expression in human peripheral blood monocytes exposed to soluble mediators known to regulate inflammation and atherogenesis. RNase protection assays showed 6- to 12-fold increases in CD36 mRNA in response to interleukin-4, monocyte colony-stimulating factor, and phorbol myristate acetate, while lipopolysaccharide and dexamethasone strongly downregulated CD36 mRNA. The downregulation of CD36 mRNA was associated with the disappearance of surface expression of CD36 antigen and loss of TSP-1 surface-binding capacity. Upregulation of CD36 mRNA was associated with a modest increase in surface antigen expression and a larger expansion of an intracellular pool of CD36. As with CD36, monocytes treated with monocyte colony-stimulating factor showed a rapid increase in TSP-1 mRNA expression. Moreover, while dexamethasone treatment decreased CD36 expression, it resulted in a rapid increase in TSP-1 mRNA, and while PMA increased CD36 mRNA, it rapidly decreased TSP-1 expression. Interferon gamma, which had no effect on CD36 mRNA, rapidly increased steady-state TSP-1 mRNA. Thus, expression of both CD36 and its ligand TSP-1 is regulated by soluble mediators, although certain mediators induce concordant changes and others discordant changes.
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PMID:Regulation of monocyte CD36 and thrombospondin-1 expression by soluble mediators. 869 41

The extreme rarity of micronucleated reticulocytes (RETs) in the peripheral blood of non-splenectomized humans has precluded facile enumeration of these cells, as well as evaluation of this endpoint as an index of cytogenetic damage. In this report, we describe a high-throughput, single-laser flow cytometric system for scoring the incidence of micronuclei (MN) in newly formed human RETs. The procedure is based on an immunochemical reagent that differentially labels the most immature fraction of RETs from mature erythrocytes based on the expression level of the transferrin receptor (also known as CD71). The resolution of four erythrocyte populations (young RETs and mature erythrocytes, with and without MN) was achieved for human blood cells treated with phycoerythrin-conjugated anti-CD71, RNase, and either SYTOX Green or SYBR Green I nucleic acid dyes. Anti-glycophorin A labeling of erythroid cells (CyChrome conjugate) was also incorporated into the staining procedure to ensure that debris or other potential artifacts did not adversely impact the analyses. Instrument calibration procedures utilizing malaria-infected rodent erythrocytes were also developed, and are described. Using this analytical system, blood samples from 10 healthy non-splenectomized human volunteers were analyzed for micronucleus frequencies with a single-laser flow cytometer. Average micronucleus frequencies in the mature and most immature fraction of RETs were 0.016 and 0.19%, respectively. Blood samples from three healthy splenectomized volunteers were also evaluated. As expected, these samples exhibited higher micronucleus frequencies in the mature subset of erythrocytes (range 0.03-0.18%). The resulting data suggest that MN can be quantified in human erythrocyte populations with a single-laser flow cytometer, and that the frequency of MN cells in the youngest reticulocyte population approaches values expected in the absence of splenic selection against MN-erythrocytes. This high throughput system is potentially important for evaluating the value of the micronucleated reticulocyte endpoint as an index of chromosome breakage and/or chromosome segregational abnormalities in human populations.
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PMID:Enumeration of micronucleated CD71-positive human reticulocytes with a single-laser flow cytometer. 1190 50

Deoxyribonucleic acid (DNA) gyrase is an important enzyme that facilitates the movement of replication and transcription complexes through DNA by creating negative supercoils ahead of the complex. Its presence in Plasmodium falciparum is now established and considered a good drug target since it is absent in the human host. The sequence of P. falciparum gyrase A subunit was analyzed for its messenger ribonucleic acid (mRNA) folding as well as target accessibility for ribozymes. The four GUC triplet sites identified at 334, 491, 1907, and 2642 nucleotide positions of the Gyrase A mRNA were also accessible to oligos by RNase H assay. Site GUC491 was optimally accessible followed by GUC1907, GUC334, and GUC2642 sites. Ribozymes were produced against all these sites and tested for their in vitro transcript cleavage potentials where RZ491 showed the maximum cleavage rate. Therefore, this ribozyme (RZ491) was chemically synthesized albeit with modifications so as to make it resistant against ribonuclease attack. The modified ribozyme retained its cleavage potential and was able to inhibit the P. falciparum parasite growth up to 49.54% and 74.77% at 20 and 30 microM ribozyme concentrations, respectively, as compared to the untreated culture. However, up to 20% and 24.32% parasite growth inhibition was observed at the same ribozyme concentrations of 20 and 30 microM when compared with control ribozyme-treated cultures. This ribozyme as well as other targets identified here can be investigated further to develop the effective chemotherapeutic agents against malaria.
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PMID:Ribozyme cleavage of Plasmodium falciparum gyrase A gene transcript affects the parasite growth. 1852 2

The traditional light microscopy has limitations for precise growth assays of malaria parasites in culture or for assessment of new compounds for antimalarial activity; the speed and high reproducibility of flow cytometry can overcome these limitations. A flow cytometric method using PicoGreen, a DNA-binding fluorochrome, was developed with optimal precision suitable for performing growth assays of low-parasitemia field isolates. In addition, intra- and inter-person reproducibility of the flow cytometric and the microscopic method were compared in order to quantitatively demonstrate the improved precision. RNase treatment contributed to the precision of the flow cytometric measurements by enhancing the signal-to-noise ratios. Coefficients of variation of the method were smaller than 10% for 0.1% or higher parasitemia samples. The intra- and inter-person coefficients of variation of the flow cytometric method were three to six times smaller than those of the microscopic method. The flow cytometric method developed in this study yielded substantially more precise results than the microscopic method, allowing determination of parasitemia levels of 0.1% or higher, with coefficients of variation smaller than 10%. Thus, the PicoGreen method could be a reliable high sensitivity assay for analysis of low parasitemia samples and might be applied to a high throughput system testing antimalarial drug activity.
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PMID:Quantitative determination of Plasmodium parasitemia by flow cytometry and microscopy. 2309 8

Sequence-specific recognition of DNA by small turn-on fluorescence probes is a promising tool for bioimaging, bioanalytical and biomedical applications. Here, the authors report a novel cell-permeable and red fluorescent hemicyanine-based thiazole coumarin (TC) probe for DNA recognition, nuclear staining and cell cycle analysis. TC exhibited strong fluorescence enhancement in the presence of DNA containing AT-base pairs, but did not fluoresce with GC sequences, single-stranded DNA, RNA and proteins. The fluorescence staining of HeLa S3 and HEK 293 cells by TC followed by DNase and RNase digestion studies depicted the selective staining of DNA in the nucleus over the cytoplasmic region. Fluorescence-activated cell sorting (FACS) analysis by flow cytometry demonstrated the potential application of TC in cell cycle analysis in HEK 293 cells. Metaphase chromosome and malaria parasite DNA imaging studies further confirmed the in vivo diagnostic and therapeutic applications of probe TC. Probe TC may find multiple applications in fluorescence spectroscopy, diagnostics, bioimaging and molecular and cell biology.
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PMID:A thiazole coumarin (TC) turn-on fluorescence probe for AT-base pair detection and multipurpose applications in different biological systems. 2525 96

Malaria is caused by a unicellular protozoan pathogen of the genus Plasmodium. Although genes represent monocistronic units that are expressed in a life cycle stage-specific manner, post-transcriptional regulation via translational repression of mRNA has been observed in parasite stages that transition from the vertebrate host to the Anopheles vector. An interesting new type of post-transcriptional control was recently discovered in Plasmodium falciparum stages that infect human erythrocytes. A subgroup of genes that were thought to be transcriptionally silent are actually transcribed but degraded immediately by an RNase II that is recruited to these gene loci. This cryptic RNA is not detectable in steady-state RNA but has been detected using nuclear run-on techniques and in mutant RNase II parasites. Nascent RNA degradation controls virulence genes expressed in a monoallelic fashion and noncoding RNAs (ncRNAs), but also a number of housekeeping-like of genes. More studies on other life cycle stages may reveal the full extent of this type of gene regulation in malaria parasites. It is tempting to speculate that RNase II-mediated gene control may exist in other eukaryotic organisms.
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PMID:RNase II: A new player enters the game. 2589 18

Malaria is a devastating illness that causes approximately 500,000 deaths annually. The malaria-causing parasite (Plasmodium genus) uses the process of translational repression to regulate its growth, development, and transmission. As poly(A)-binding proteins (PABP) have been identified as critical components of RNA metabolism and translational repression in model eukaryotes and in Plasmodium, we have identified and investigated two PABPs in Plasmodium yoelii, PyPABP1 and PyPABP2. In contrast to most single-celled eukaryotes, Plasmodium closely resembles metazoans and encodes both a nuclear PABP and a cytosolic PABP; here, we provide multiple lines of evidence in support of this observation. The conserved domain architectures of PyPABP1 and PyPABP2 resemble those of yeast and metazoans, while multiple independent binding assays demonstrated their ability to bind very strongly and specifically to poly(A) sequences. Interestingly, we also observed that purified PyPABP1 forms homopolymeric chains despite exhaustive RNase treatment in vitro. Finally, we show by indirect immunofluorescence assays (IFAs) that PyPABP1 and PyPABP2 are cytoplasm- and nucleus-associated PABPs during the blood stages of the life cycle. Surprisingly, however, PyPABP1 was instead observed to also be localized on the surface of transmitted salivary gland sporozoites and to be deposited in trails when parasites glide on a substrate. This is the third RNA-binding protein verified to be found on the sporozoite surface, and the data may point to an unappreciated RNA-centered interface between the host and parasite. IMPORTANCE Malaria remains one of the great global health problems. The parasite that causes malaria (Plasmodium genus) relies upon exquisite control of its transmission between vertebrate hosts and mosquitoes. One crucial way that it does so is by proactively producing mRNAs needed to establish the new infection but by silencing and storing them until they are needed. One key protein in this process of translational repression in model eukaryotes is poly(A)-binding protein (PABP). Here we have shown that Plasmodium yoelii utilizes both a nuclear PABP and a cytosolic PABP, both of which bind specifically to polyadenylated RNA sequences. Moreover, we find that the cytosolic PABP forms chains in vitro, consistent with its appreciated role in coating the poly(A) tails of mRNA. Finally, we have also verified that, surprisingly, the cytosolic PABP is found on the surface of Plasmodium sporozoites. Taking the data together, we propose that Plasmodium utilizes a more metazoan-like strategy for RNA metabolism using specialized PABPs.
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PMID:Nuclear, Cytosolic, and Surface-Localized Poly(A)-Binding Proteins of Plasmodium yoelii. 2935 80

The malaria parasite Plasmodium and other apicomplexans such as Toxoplasma evolved from photosynthetic organisms and contain an essential, remnant plastid termed the apicoplast. Transcription of the apicoplast genome is polycistronic with extensive RNA processing. Yet little is known about the mechanism of apicoplast RNA processing. In plants, chloroplast RNA processing is controlled by multiple pentatricopeptide repeat (PPR) proteins. Here, we identify the single apicoplast PPR protein, PPR1. We show that the protein is essential and that it binds to RNA motifs corresponding with previously characterized processing sites. Additionally, PPR1 shields RNA transcripts from ribonuclease degradation. This is the first characterization of a PPR protein from a nonphotosynthetic plastid.
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PMID:An essential pentatricopeptide repeat protein in the apicomplexan remnant chloroplast. 3145 37

The heterochromatin environment plays a central role in silencing genes associated with the malaria parasite's development, survival in the host, and transmission to the mosquito vector. However, the underlying mechanism regulating the dynamic chromatin structure is not understood yet. Here, we have uncovered that Plasmodium falciparum Rrp6, an orthologue of eukaryotic RNA exosome-associated RNase, controls the silencing of heterochromatic genes. PfRrp6 knockdown disrupted the singular expression of the GC-rich ncRNA RUF6 family, a known critical regulator of virulence gene expression, through the stabilization of the nascent transcripts. Mechanistic investigation showed that the accumulation of the multiple RUF6 ncRNAs triggered local chromatin remodeling in situ, which activated their adjacent var genes. Strikingly, chromatin isolation by RNA purification analysis (ChIRP-seq) revealed that a remarkable RUF6 ncRNA had interacted with distal heterochromatin regions directly and stimulated a global derepression effect on heterochromatic genes, including all variant gene families and the sexual commitment-associated regulator ap2-g gene. Collectively, Rrp6 appears to conduct the epigenetic surveillance of heterochromatic gene expression through controlling RUF6 levels, thereby securing antigenic variation and sexual commitment of malaria parasites during the infection of the host.IMPORTANCE Malaria remains a major public health and economic burden. The heterochromatin environment controls the silencing of genes associated with the fate of malaria parasites. Previous studies have demonstrated that a group of GC-rich ncRNAs (RUF6) is associated with the mutually exclusive expression of var genes, but the underlying mechanisms remain elusive. Here, through a series of genetic manipulation and genome-wide multiomics analysis, we have identified the plasmodial orthologue of RNA exosome-associated Rrp6 as an upstream regulator of RUF6 expression and revealed that the dysregulation of RUF6 upon Rrp6 knockdown triggered local chromatin alteration, thereby activating most heterochromatic genes via direct interaction of RUF6 and distal gene loci. This finding not only uncovered the in-depth mechanism of RUF6-mediated regulation of heterochromatic genes but also identified Rrp6 as a novel regulator of gene expression in human malaria parasites, which provides a new target for developing intervention strategies against malaria.
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PMID:Rrp6 Regulates Heterochromatic Gene Silencing via ncRNA RUF6 Decay in Malaria Parasites. 3248 61


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