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Background
The family Tymoviridae comprises three plant virus genera, including Tymovirus, Marafivirus, and Maculavirus, which are found in most parts of the world and cause severe agricultural losses. We describe a putatively novel member of the family Tymoviridae, which is isolated from mosquitoes (Culex spp.), referred to as CuTLV.Methods and Results
The CuTLV was isolated by cell culture, which replicates and causes cytopathic effects in Aedes albopictus C6/36 cells, but not in mammalian BHK-21 or Vero cells. The complete 6471 nucleotide sequence of CuTLV was determined. The genome of CuTLV is predicted to contain three open reading frames (ORFs). The largest ORF1 is 5307 nucleotides (nt) in length and encodes a putative polypeptide of 1769 amino acids (aa), which contains the conserved motifs for the methyltransferase (MTR), Tymovirus endopeptidase (PRO), helicase (HEL), and RNA-dependent RNA polymerase (RdRp) of the replication-associated proteins (RPs) of positive-stranded RNA viruses. In contrast, the ORF1 sequence does not contain the so-called “tymobox” or “marafibox”, the conserved subgenomic RNA promoter present in all tymoviruses or marafiviruses, respectively. ORF2 and ORF3 putatively encode a 248-aa coat protein (CP) and a proline-rich 149-aa polypeptide. The whole genomic nucleotide identity of CuTLV with other members of family Tymoviridae ranged from 46.2% (ChiYMV) to 52.4% (GFkV). Phylogenetic analysis based on the putative RP and CP genes of CuTLV demonstrated that the virus is most closely related to viruses in the genus Maculavirus.Conclusions
The CuTLV is a novel virus related to members of the family Tymoviridae, with molecular characters that are distinct from those of tymoviruses, marafiviruses, and other maculaviruses or macula-like viruses. This is the first report of the isolation of a Tymoviridae-like virus from mosquitoes. Further investigations are required to clarify the origin, replication strategy, and the public health or agricultural importance of the CuTLV. 相似文献3.
Background
The Gadd45 proteins play important roles in growth control, maintenance of genomic stability, DNA repair, and apoptosis. Recently, Gadd45 proteins have also been implicated in epigenetic gene regulation by promoting active DNA demethylation. Gadd45 proteins have sequence homology with the L7Ae/L30e/S12e RNA binding superfamily of ribosomal proteins, which raises the question if they may interact directly with nucleic acids.Principal Findings
Here we show that Gadd45a binds RNA but not single- or double stranded DNA or methylated DNA in vitro. Sucrose density gradient centrifugation experiments demonstrate that Gadd45a is present in high molecular weight particles, which are RNase sensitive. Gadd45a displays RNase-sensitive colocalization in nuclear speckles with the RNA helicase p68 and the RNA binding protein SC35. A K45A point mutation defective in RNA binding was still active in DNA demethylation. This suggests that RNA binding is not absolutely essential for demethylation of an artificial substrate. A point mutation at G39 impared RNA binding, nuclear speckle localization and DNA demethylation, emphasizing its relevance for Gadd45a function.Significance
The results implicate RNA in Gadd45a function and suggest that Gadd45a is associated with a ribonucleoprotein particle. 相似文献4.
Mielecki D Zugaj DŁ Muszewska A Piwowarski J Chojnacka A Mielecki M Nieminuszczy J Grynberg M Grzesiuk E 《PloS one》2012,7(1):e30588
Background
ALKBH proteins, the homologs of Escherichia coli AlkB dioxygenase, constitute a direct, single-protein repair system, protecting cellular DNA and RNA against the cytotoxic and mutagenic activity of alkylating agents, chemicals significantly contributing to tumor formation and used in cancer therapy. In silico analysis and in vivo studies have shown the existence of AlkB homologs in almost all organisms. Nine AlkB homologs (ALKBH1–8 and FTO) have been identified in humans. High ALKBH levels have been found to encourage tumor development, questioning the use of alkylating agents in chemotherapy. The aim of this work was to assign biological significance to multiple AlkB homologs by characterizing their activity in the repair of nucleic acids in prokaryotes and their subcellular localization in eukaryotes.Methodology and Findings
Bioinformatic analysis of protein sequence databases identified 1943 AlkB sequences with eight new AlkB subfamilies. Since Cyanobacteria and Arabidopsis thaliana contain multiple AlkB homologs, they were selected as model organisms for in vivo research. Using E. coli alkB − mutant and plasmids expressing cyanobacterial AlkBs, we studied the repair of methyl methanesulfonate (MMS) and chloroacetaldehyde (CAA) induced lesions in ssDNA, ssRNA, and genomic DNA. On the basis of GFP fusions, we investigated the subcellular localization of ALKBHs in A. thaliana and established its mostly nucleo-cytoplasmic distribution. Some of the ALKBH proteins were found to change their localization upon MMS treatment.Conclusions
Our in vivo studies showed highly specific activity of cyanobacterial AlkB proteins towards lesions and nucleic acid type. Subcellular localization and translocation of ALKBHs in A. thaliana indicates a possible role for these proteins in the repair of alkyl lesions. We hypothesize that the multiplicity of ALKBHs is due to their involvement in the metabolism of nucleo-protein complexes; we find their repair by ALKBH proteins to be economical and effective alternative to degradation and de novo synthesis. 相似文献5.
Background
Bluetongue virus (BTV) protein, VP1, is known to possess an intrinsic polymerase function, unlike rotavirus VP1, which requires the capsid protein VP2 for its catalytic activity. However, compared with the polymerases of other members of the Reoviridae family, BTV VP1 has not been characterized in detail.Methods and Findings
Using an in vitro polymerase assay system, we demonstrated that BTV VP1 could synthesize the ten dsRNAs simultaneously from BTV core-derived ssRNA templates in a single in vitro reaction as well as genomic dsRNA segments from rotavirus core-derived ssRNA templates that possess no sequence similarity with BTV. In contrast, dsRNAs were not synthesized from non-viral ssRNA templates by VP1, unless they were fused with specific BTV sequences. Further, we showed that synthesis of dsRNAs from capped ssRNA templates was significantly higher than that from uncapped ssRNA templates and the addition of dinucleotides enhanced activity as long as the last base of the dinucleotide complemented the 3′ -terminal nucleotide of the ssRNA template.Conclusions
We showed that the polymerase activity was stimulated by two different factors: cap structure, likely due to allosteric effect, and dinucleotides due to priming. Our results also suggested the possible presence of cis-acting elements shared by ssRNAs in the members of family Reoviridae. 相似文献6.
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Background
RNAi technology is widely used to downregulate specific gene products. Investigating the phenotype induced by downregulation of gene products provides essential information about the function of the specific gene of interest. When RNAi is applied in Drosophila melanogaster or Caenorhabditis elegans, often large dsRNAs are used. One of the drawbacks of RNAi technology is that unwanted gene products with sequence similarity to the gene of interest can be down regulated too. To verify the outcome of an RNAi experiment and to avoid these unwanted off-target effects, an additional non-overlapping dsRNA can be used to down-regulate the same gene. However it has never been tested whether this approach is sufficient to reduce the risk of off-targets.Methodology
We created a novel tool to analyse the occurance of off-target effects in Drosophila and we analyzed 99 randomly chosen genes.Principal Findings
Here we show that nearly all genes contain non-overlapping internal sequences that do show overlap in a common off-target gene.Conclusion
Based on our in silico findings, off-target effects should not be ignored and our presented on-line tool enables the identification of two RNA interference constructs, free of overlapping off-targets, from any gene of interest. 相似文献8.
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Jan Felix Drexler Victor Max Corman Florian Gloza-Rausch Antje Seebens Augustina Annan Anne Ipsen Thomas Kruppa Marcel A. Müller Elisabeth K. V. Kalko Yaw Adu-Sarkodie Samuel Oppong Christian Drosten 《PloS one》2009,4(7)
Background
Henipaviruses (Hendra and Nipah virus) are highly pathogenic members of the family Paramyxoviridae. Fruit-eating bats of the Pteropus genus have been suggested as their natural reservoir. Human Henipavirus infections have been reported in a region extending from Australia via Malaysia into Bangladesh, compatible with the geographic range of Pteropus. These bats do not occur in continental Africa, but a whole range of other fruit bats is encountered. One of the most abundant is Eidolon helvum, the African Straw-coloured fruit bat.Methodology/Principal Findings
Feces from E. helvum roosting in an urban setting in Kumasi/Ghana were tested for Henipavirus RNA. Sequences of three novel viruses in phylogenetic relationship to known Henipaviruses were detected. Virus RNA concentrations in feces were low.Conclusions/Significance
The finding of novel putative Henipaviruses outside Australia and Asia contributes a significant extension of the region of potential endemicity of one of the most pathogenic virus genera known in humans. 相似文献10.
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Background
In silico models have recently been created in order to predict which genetic variants are more likely to contribute to the risk of a complex trait given their functional characteristics. However, there has been no comprehensive review as to which type of predictive accuracy measures and data visualization techniques are most useful for assessing these models.Methods
We assessed the performance of the models for predicting risk using various methodologies, some of which include: receiver operating characteristic (ROC) curves, histograms of classification probability, and the novel use of the quantile-quantile plot. These measures have variable interpretability depending on factors such as whether the dataset is balanced in terms of numbers of genetic variants classified as risk variants versus those that are not.Results
We conclude that the area under the curve (AUC) is a suitable starting place, and for models with similar AUCs, violin plots are particularly useful for examining the distribution of the risk scores.Electronic supplementary material
The online version of this article (doi:10.1186/s12864-015-1616-z) contains supplementary material, which is available to authorized users. 相似文献12.
Guillermo Pastor-Palacios Elisa Azuara-Liceaga Luis G. Brieba 《PLoS neglected tropical diseases》2010,4(8)
Background
Eukaryotic family A DNA polymerases are involved in mitochondrial DNA replication or translesion DNA synthesis. Here, we present evidence that the sole family A DNA polymerase from the parasite protozoan E. histolytica (EhDNApolA) localizes to the nucleus and that its biochemical properties indicate that this DNA polymerase may be involved in translesion DNA synthesis.Methodology and Results
EhDNApolA is the sole family A DNA polymerase in E. histolytica. An in silico analysis places family A DNA polymerases from the genus Entamoeba in a separate branch of a family A DNA polymerases phylogenetic tree. Biochemical studies of a purified recombinant EhDNApolA demonstrated that this polymerase is active in primer elongation, is poorly processive, displays moderate strand displacement, and does not contain 3′–5′ exonuclease or editing activity. Importantly, EhDNApolA bypasses thymine glycol lesions with high fidelity, and confocal microscopy demonstrates that this polymerase is translocated into the nucleus. These data suggest a putative role of EhDNApolA in translesion DNA synthesis in E. histolytica.Conclusion
This is the first report of the biochemical characterization of a DNA polymerase from E. histolytica. EhDNApolA is a family A DNA polymerase that is grouped into a new subfamily of DNA polymerases with translesion DNA synthesis capabilities similar to DNA polymerases from subfamily ν. 相似文献13.
Laha T Loukas A Wattanasatitarpa S Somprakhon J Kewgrai N Sithithaworn P Kaewkes S Mitreva M Brindley PJ 《PLoS neglected tropical diseases》2007,1(1):e35
Background
An enhanced understanding of the hookworm genome and its resident mobile genetic elements should facilitate understanding of the genome evolution, genome organization, possibly host-parasite co-evolution and horizontal gene transfer, and from a practical perspective, development of transposon-based transgenesis for hookworms and other parasitic nematodes.Methodology/Principal Findings
A novel mariner-like element (MLE) was characterized from the genome of the dog hookworm, Ancylostoma caninum, and termed bandit. The consensus sequence of the bandit transposon was 1,285 base pairs (bp) in length. The new transposon was flanked by perfect terminal inverted repeats of 32 nucleotides in length with a common target site duplication TA, and it encoded an open reading frame (ORF) of 342 deduced amino acid residues. Phylogenetic comparisons confirmed that the ORF encoded a mariner-like transposase, which included conserved catalytic domains, and that the bandit transposon belonged to the cecropia subfamily of MLEs. The phylogenetic analysis also indicated that the Hsmar1 transposon from humans was the closest known relative of bandit, and that bandit and Hsmar1 constituted a clade discrete from the Tc1 subfamily of MLEs from the nematode Caenorhabditis elegans. Moreover, homology models based on the crystal structure of Mos1 from Drosophila mauritiana revealed closer identity in active site residues of the catalytic domain including Ser281, Lys289 and Asp293 between bandit and Hsmar1 than between Mos1 and either bandit or Hsmar1. The entire bandit ORF was amplified from genomic DNA and a fragment of the bandit ORF was amplified from RNA, indicating that this transposon is actively transcribed in hookworms.Conclusions/Significance
A mariner-like transposon termed bandit has colonized the genome of the hookworm A. caninum. Although MLEs exhibit a broad host range, and are identified in other nematodes, the closest phylogenetic relative of bandit is the Hsmar1 element of humans. This surprising finding suggests that bandit was transferred horizontally between hookworm parasites and their mammalian hosts. 相似文献14.
Janett Fischer Stephan B?hm Tobias Müller Heiko Witt Christoph Sarrazin Simone Susser Pascal Migaud Eckart Schott Graeme Stewart Annika Brodzinski Balazs Fül?p Florian van B?mmel Jacob George Thomas Berg 《PloS one》2013,8(10)
Background & Aims
Genetic variations near the interferon lambda 3 gene (IFNL3, IL28B) are the most powerful predictors for sustained virologic response (SVR) in patients with chronic hepatitis C virus (HCV) infection, compared to other biochemical or histological baseline parameters. We evaluated whether the interplay of both IFNL3 polymorphisms rs12979860 and rs8099917 together with non-genetic clinical factors contributes to the predictive role of these genetic variants.Methods
The cohort comprised 1,402 patients of European descent with chronic HCV type 1 infection. 1,298 patients received interferon-based antiviral therapy, and 719 (55%) achieved SVR. The IFNL3 polymorphisms were genotyped by polymerase chain reaction and melting curve analysis.Results
A significant correlation was found between the IFNL3 polymorphisms and biochemical as well as virologic predictors of treatment outcome such as ALT, GGT, cholesterol, and HCV RNA levels. In multivariate regression analysis, IFLN3 SNPs, HCV RNA levels, and the GGT/ALT ratio were independent predictors of SVR. Dependent on the GGT/ALT ratio and on the HCV RNA concentration, significant variations in the likelihood for achieving SVR were observed in both, carriers of the responder as well as non-responder alleles.Conclusions
Our data support a clear association between IFNL3 genotypes and baseline parameters known to impact interferon responsiveness. Improved treatment outcome prediction was achieved when these predictors were considered in combination with the IFNL3 genotype. 相似文献15.
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Background
Lactate dehydrogenase-elevating virus (LDV) is a natural infectious agent of mice. Like several other viruses, LDV causes widespread and very rapid but transient activation of both B cells and T cells in lymphoid tissues and the blood. The mechanism of this activation has not been fully described and is the focus of the current studies.Principal Findings
A known inducer of early lymphocyte activation is IFNα, a cytokine strongly induced by LDV infection. Neutralization of IFNα in the plasma from infected mice ablated its ability to activate lymphocytes in vitro. Since the primary source of virus-induced IFNα in vivo is often plasmacytoid dendritic cells (pDC''s), we depleted these cells prior to LDV infection and tested for lymphocyte activation. Depletion of pDC''s in vivo eradicated both the LDV-induced IFNα response and lymphocyte activation. A primary receptor in pDC''s for single stranded RNA viruses such as LDV is the toll-like receptor 7 (TLR7) pattern recognition receptor. Infection of TLR7-knockout mice revealed that both the IFNα response and lymphocyte activation were dependent on TLR7 signaling in vivo. Interestingly, virus levels in both TLR7 knockout mice and pDC-depleted mice were indistinguishable from controls indicating that LDV is largely resistant to the systemic IFNα response.Conclusion
Results indicate that LDV-induced activation of lymphocytes is due to recognition of LDV nucleic acid by TLR7 pattern recognition receptors in pDC''s that respond with a lymphocyte-inducing IFNα response. 相似文献17.
Background
The order Rickettsiales comprises Gram-negative obligate intracellular bacteria (also called rickettsias) that are mainly associated with arthropod hosts. This group is medically important because it contains human-pathogenic species that cause dangerous diseases. Until now, there has been no report of non-phagotrophic photosynthetic eukaryotes, such as green plants, harboring rickettsias.Methodology/Principal Findings
We examined the bacterial endosymbionts of two freshwater volvocalean green algae: unicellular Carteria cerasiformis and colonial Pleodorina japonica. Epifluorescence microscopy using 4′-6-deamidino-2-phenylindole staining revealed the presence of endosymbionts in all C. cerasiformis NIES-425 cells, and demonstrated a positive correlation between host cell size and the number of endosymbionts. Strains both containing and lacking endosymbionts of C. cerasiformis (NIES-425 and NIES-424) showed a >10-fold increase in cell number and typical sigmoid growth curves over 192 h. A phylogenetic analysis of 16 S ribosomal (r)RNA gene sequences from the endosymbionts of C. cerasiformis and P. japonica demonstrated that they formed a robust clade (hydra group) with endosymbionts of various non-arthropod hosts within the family Rickettsiaceae. There were significantly fewer differences in the 16 S rRNA sequences of the rickettsiacean endosymbionts between C. cerasiformis and P. japonica than in the chloroplast 16 S rRNA or 18 S rRNA of the host volvocalean cells. Fluorescence in situ hybridization demonstrated the existence of the rickettsiacean endosymbionts in the cytoplasm of two volvocalean species.Conclusions/Significance
The rickettsiacean endosymbionts are likely not harmful to their volvocalean hosts and may have been recently transmitted from other non-arthropod organisms. Because rickettsias are the closest relatives of mitochondria, incipient stages of mitochondrial endosymbiosis may be deduced using both strains with and without C. cerasiformis endosymbionts. 相似文献18.
Seiji Shiota Rumiko Suzuki Yuichi Matsuo Muhammad Miftahussurur Trang Thu Huyen Tran Tran Thanh Binh Yoshio Yamaoka 《PloS one》2014,9(8)
Background
A recent report has shown that the phylogenetic origin of Helicobacter pylori based on multi-locus sequence typing (MLST) was significantly associated with the severity of gastritis in Colombia. However, the potential relationship between phylogenetic origin and clinical outcomes was not examined in that study. If the phylogenetic origin rather than virulence factors were truly associated with clinical outcomes, identifying a population at high risk for gastric cancer in Colombia would be relatively straightforward. In this study, we examined the phylogenetic origins of strains from gastric cancer and duodenal ulcer patients living in Bogota, Colombia.Methods
We included 35 gastric cancer patients and 31 duodenal ulcer patients, which are considered the variant outcomes. The genotypes of cagA and vacA were determined by polymerase chain reaction. The genealogy of these Colombian strains was analyzed by MLST. Bacterial population structure was analyzed using STRUCTURE software.Results
H. pylori strains from gastric cancer and duodenal ulcer patients were scattered in the phylogenetic tree; thus, we did not detect any difference in phylogenetic distribution between gastric cancer and duodenal ulcer strains in the hpEurope group in Colombia. Sixty-six strains, with one exception, were classified as hpEurope irrespective of the cagA and vacA genotypes, and type of disease. STRUCTURE analysis revealed that Colombian hpEurope strains have a phylogenetic connection to Spanish strains.Conclusions
Our study showed that a phylogeographic origin determined by MLST was insufficient for distinguishing between gastric cancer and duodenal ulcer risk among hpEurope strains in the Andean region in Colombia. Our analysis also suggests that hpEurope strains in Colombia were primarily introduced by Spanish immigrants. 相似文献19.
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Raúl Teruel Irene Martínez-Martínez José A Guerrero Rocío González-Conejero María E de la Morena-Barrio Salam Salloum-Asfar Ana B Arroyo Sonia águila Nuria García-Barberá Antonia Mi?ano Vicente Vicente Javier Corral Constantino Martínez 《Journal of biomedical science》2013,20(1):29