Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

By combining molecular-biological techniques with our increased understanding of the effect of gene sequence modification on viral function, yellow fever 17D, a positive-strand RNA virus vaccine, has been manipulated to induce a protective immune response against viruses of the same family (e.g. Japanese encephalitis and dengue viruses). Triggered by the emergence of West Nile virus infections in the New World afflicting humans, horses and birds, the success of this recombinant technology has prompted the rapid development of a live-virus attenuated candidate vaccine against West Nile virus.
Trends Mol Med 2001 Aug
PMID:Yellow fever vector live-virus vaccines: West Nile virus vaccine development. 1151 95

Mosquito-vectored diseases such as yellow fever and dengue fever continue to have a substantial impact on human populations world-wide. Novel strategies for control of these mosquito vectored diseases can arise through the development of reliable systems for genetic manipulation of the insect vector. A piggyBac vector marked with the Drosophila melanogaster cinnabar (cn) gene was used to transform the white-eyed khw strain of Aedes aegypti. Microinjection of preblastoderm embryos resulted in four families of cinnabar transformed insects. An overall transformation frequency of 4%, with a range of 0% to as high as 13% for individual experiments, was achieved when using a heat-shock induced transposase providing helper plasmid. Southern hybridizations indicated multiple insertion events in three of four transgenic lines, while the presence of duplicated target TTAA sites at either ends of individual insertions confirmed characteristic piggyBac transposition events in these three transgenic lines. The transgenic phenotype has remained stable for more than twenty generations. The transformations effected using the piggyBac element establish the potential of this element as a germ-line transformation vector for Aedine mosquitoes.
Insect Mol Biol 2002 Apr
PMID:Germ line transformation of the yellow fever mosquito, Aedes aegypti, mediated by transpositional insertion of a piggyBac vector. 1196 78

Mosquitoes transmit numerous viral pathogens to humans including dengue virus which affects approximately 50 million individuals per year. Inhibition of viral gene expression within an insect host could be used to block virus replication and subsequent transmission of the pathogen to humans. A naturally occurring gene silencing mechanism triggered by double-stranded RNA (dsRNA), RNA interference (RNAi), has recently been described in a number of species including Drosophila. To ascertain if dsRNA-triggered RNAi is present in mosquito cells, we used Aedes albopictus C6/36 cells, and to investigate the feasibility of blocking viral gene expression and replication, we used two mosquito-borne viruses, Semliki Forest virus (SFV) and the serotype 1 dengue virus (DEN1). We demonstrate that dsRNA can specifically inhibit transgene expression in C6/36 cells from both plasmid and SFV replicons and can significantly modify the kinetics of DEN1 RNA and virus replication. The inhibition mediated by dsRNA was sequence-specific and either equal or superior to that induced by antisense single-stranded RNA (ssRNA). This study demonstrates dsRNA-triggered inhibition of gene expression and virus replication in mosquito cells and suggests that this mechanism could be used to block pathogen replication within an insect host and, thus, block disease transmission.
Mol Ther 2002 Aug
PMID:Inhibition of viral gene expression and replication in mosquito cells by dsRNA-triggered RNA interference. 1216 Nov 91

Diseases caused by arthropod-borne viruses are increasingly significant public health problems, and novel methods are needed to control pathogen transmission. The hypothesis underlying the research described here is that genetic manipulation of Aedes aegypti mosquitoes can profoundly and permanently reduce their competence to transmit dengue viruses to human hosts. Recent key findings now allow us to test the genetic control hypothesis. We have identified viral genome-derived RNA segments that can be expressed in mosquito midguts and salivary glands to ablate homologous virus replication and transmission. We have demonstrated that both transient and heritable expression of virus-derived effector RNAs in cultured mosquito cells can silence virus replication, and have characterized the mechanism of RNA-mediated resistance. We are now developing virus-resistant mosquito lines by transformation with transposable elements that express effector RNAs from mosquito-active promoters.
Insect Biochem Mol Biol 2002 Oct
PMID:Developing arbovirus resistance in mosquitoes. 1222 24

The NS2B-NS3(pro) polyprotein segment from the dengue virus serotype 2 strain 16681 was purified from overexpressing E. coli by metal chelate affinity chromatography and gel filtration. Enzymatic activity of the refolded NS2B-NS3(pro) protease complex was determined in vitro with dansyl-labeled peptide substrates, based upon native dengue virus type 2 cleavage sites. The 12mer substrate peptides and the cleavage products could be separated by reversed-phase HPLC, and were identified by UV and fluorescence detection. All of the peptide substrates (representing the DEN polyprotein junction sequences at the NS2A/NS2B, NS2B/NS3, NS3/NS4A and NS4B/NS5 sites) were cleaved by the recombinant protease NS2B-NS3(pro). No cleavage was observed with an enzymatically inactive S135A mutant of the NS3 protein, or with a modified substrate peptide of the NS3/NS4A polyprotein site that contained a K2093A substitution. Enzymatic activity was dependent on the salt concentration. A 50% decrease of activity was observed in the presence of 0.1 M sodium chloride. Our results show that the NS3 protease activity of the refolded NS2BNS3(pro) protein can be assayed in vitro with high specificity by using cleavage-junction derived peptide substrates.
J Biochem Mol Biol 2002 Mar 31
PMID:In vitro determination of dengue virus type 2 NS2B-NS3 protease activity with fluorescent peptide substrates. 1229 31

Isoenzyme variation was assessed in 79 mosquito samples of Aedes aegypti, and susceptibility to a dengue 2 virus strain was evaluated in 83 samples. Analysis of FST values, differentiation indexes, and geographic distances separating populations revealed that genetic differences between populations depended on the species' history of migration and colonization. Three major clusters were identified: (1). the sylvan form, Ae. ae. formosus, from West Africa and some islands in the Indian Ocean; (2). the domestic form, Ae. ae. aegypti, from Southeast Asia and South America; and (3). Ae. ae. aegypti populations from the South Pacific islands. Two groups were identified on the basis of susceptibility to the dengue virus: (1). populations with high infection rates, mostly the Ae. ae. aegypti form, and (2). mosquitoes with lower infection rates, specifically Ae. ae. formosus. Other evolutionary and epidemiological implications of the genetic variability of Ae. aegypti are also discussed.
J Mol Evol 2002 Dec
PMID:Geographic genetic variation in populations of the dengue virus vector Aedes aegypti. 1248 24

Dengue is often referred to as an emerging disease because of the rapid increases in incidence and prevalence that have been observed in recent decades. To understand the rate at which genetic diversification occurs in dengue virus and to infer the time-scale of its evolution, we employed a maximum likelihood method that uses information about times of virus sampling to estimate the rate of molecular evolution in a large number of viral envelope (E) gene sequences and to place bounds around the dates of appearance of all serotypes and specific genotypes. Our analysis reveals that dengue virus generally evolves according to a molecular clock, although some serotype-specific and genotype-specific rate differences were observed, and that its origin is more recent than previously suggested, with the virus appearing approximately 1,000 years ago. Furthermore, we estimate that the zoonotic transfer of dengue from sylvatic (monkey) to sustained human transmission occurred between 125 and 320 years ago, that the current global genetic diversity in the four serotypes of dengue virus only appeared during the past century, and that the recent rise in genetic diversity can be loosely correlated both to human activities such as population growth, urbanization, and mass transport and to the emergence of dengue hemorrhagic fever as a major disease problem.
Mol Biol Evol 2003 Jan
PMID:Inferring the rate and time-scale of dengue virus evolution. 1251 14

The possibility of obtaining, from any antibody, a fluorescent conjugate which responds to the binding of the antigen by a variation of fluorescence, would be of great interest in the micro- and nano-analytical sciences. This possibility was explored with antibody mAb4E11, which is directed against the dengue virus and for which no structural data is available. Three rules of design were developed to identify residues of the antibody to which a fluorophore could be chemically coupled, after changing them to cysteine by mutagenesis. (i) The target residue belonged to the hypervariable loops of the antibody. (ii) It was adjacent, along the amino acid sequence of the antibody, to a residue which was functionally important for the interaction with the antigen. (iii) It was not important in itself for the interaction with the antigen. Eight conjugates between a single chain variable fragment of mAb4E11 and an environment-sensitive fluorophore were constructed. Three of them showed an increase in their fluorescence intensity by 1.5-2.8-fold on antigen binding, without loss of affinity. This increase allowed the titration of the antigen in serum above a threshold concentration of 10nM. Experiments of quenching with potassium iodide suggested that the fluorescence variation was due to a shielding of the fluorescent group from the solvent by the binding of the antigen, and that therefore its mechanism is general.
J Mol Biol 2003 Feb 07
PMID:Deriving topological constraints from functional data for the design of reagentless fluorescent immunosensors. 1254 99

In the last four decades the incidence of dengue fever has increased 30-fold worldwide, and over half the world's population is now threatened with infection from one or more of four co-circulating viral serotypes (DEN-1 through DEN-4). To determine the role of viral molecular evolution in emergent disease dynamics, we sequenced 40% of the genome of 82 DEN-4 isolates collected from Puerto Rico over the 20 years since the onset of endemic dengue on the island. Isolates were derived from years with varying levels of DEN-4 prevalence. Over our sampling period there were marked evolutionary shifts in DEN-4 viral populations circulating in Puerto Rico; viral lineages were temporally clustered and the most common genotype at a particular sampling time often arose from a previously rare lineage. Expressed changes in structural genes did not appear to drive this lineage turnover, even though these regions include primary determinants of viral antigenic properties. Instead, recent dengue evolution can be attributed in part to positive selection on the nonstructural gene 2A (NS2A), whose functions may include replication efficiency and antigenicity. During the latest and most severe DEN-4 epidemic in Puerto Rico, in 1998, viruses were distinguished by three amino acid changes in NS2A that were fixed far faster than expected by drift alone. Our study therefore demonstrates viral genetic turnover within a focal population and the potential importance of adaptive evolution in viral epidemic expansion.
Mol Biol Evol 2003 Oct
PMID:Selection-driven evolution of emergent dengue virus. 1283 29

The epidemic of West Nile virus (WNV) in the USA in 2002 represents the largest outbreak of meningoencephalitis in the Western Hemisphere ever reported. Besides natural transmission by mosquitoes, five new modes of WNV transmission to humans have been reported: blood transfusion, organ transplantation, transplacental transmission, breastfeeding and laboratory-acquired infection. The recognition of these new transmission routes has made the development of rapid and accurate serological diagnosis of WNV infection a public health priority. In this article, the current serologic assays for WNV diagnosis are reviewed, including immunoglobulin M antibody-capture ELISA, immunoglobulin G ELISA, indirect fluorescent antibody tests, hemagglutination inhibition tests and plaque reduction neutralization tests. The recently developed immunoassays that use purified recombinant envelope and nonstructural protein 5 of WNV as antigens are also reviewed. The nonstructural protein 5 protein-based assay can reliably discriminate between WNV and dengue or St. Louis encephalitis virus, as well as between natural WNV infection and flavivirus vaccination.
Expert Rev Mol Diagn 2003 Nov
PMID:Serologic diagnosis of West Nile virus infection. 1462 1


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