Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.48 (transcriptase)
9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Haemophilus parasuis is an important opportunistic pathogen in swine of high health status, but to date no proven virulence factors have been described. As virulence factors are known to be regulated during disease, the objective of this study was to identify genes of a virulent serovar 5 strain with altered expression after iron restriction or in the presence of porcine cerebrospinal fluid (CSF), conditions that reflect in vivo growth conditions. Using differential-display reverse-transcriptase-mediated polymerase chain reaction, we found that homologues of genes encoding fructose bisphosphate aldolase (fba), adenylosuccinate synthetase (purA), 2',3'-cyclic nucleotide phosphodiesterase (cpdB), lipoprotein signal peptidase (lspA), pyrophosphate reductase (lytB), superoxide dismutase (sodC), tyrosyl t-RNA synthetase (tyrS), cysteine synthetase (cysK), an unknown protein, and a homologue of a hydrolase of the haloacid dehydrogenase superfamily were upregulated in response to iron restriction. In addition, the purA, cpdB, lspA, lytB, and sodC homologues, cDNAs homologous with a Na+/alanine symporter, fatty acid ligase (fadD), diadenosine tetraphosphatase (apaH), and an unknown protein were upregulated in response to CSF. In screening for the presence of these differentially expressed genes to assess their usefulness as diagnostic markers of high virulence potential, we detected homologues of all of these genes in all of the reference strains of the 15 established serovars. The hydrolase homologue, however, was expressed only in representative H. parasuis strains associated with a high virulence potential, suggesting that this enzyme may play a role in pathogenesis.
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PMID:Differential expression of Haemophilus parasuis genes in response to iron restriction and cerebrospinal fluid. 1769 92

We previously demonstrated that a high dose of tacrolimus (1 mumol/L) induced expression of matrix metalloproteinase (MMP) proteins in human cultured gingival fibroblasts, suggesting a molecular mechanism maintaining gingival collagen homeostasis in tacrolimus-treated patients. Herein we have analyzed whether the effect on collagen turnover might be influenced by a therapeutic tacrolimus dose. Human gingival fibroblasts were incubated for 72 hours with 10 nmol/L, 100 nmol/L, and 1 mumol/L tacrolimus, or left untreated (CT). Collagen type I and III (COL-I, COL-III), lysyl hydroxylase 2b (LH2b), MMP-1 and -2, tissue inhibitor of MMP-1 and transforming growth factor-beta1 (TGF-beta1) mRNA levels were assayed by reverse-transcriptase polymerase chain reaction, collagen protein levels by dot blot, and MMP activity by sodium dodecyl sulfate zymography. Tacrolimus did not affect COL-I, COL-III, or MMP gene expression, while LH2b and TGF-beta1 tended to be down-regulated after 1 mumol/L FK506. Conversely, protein levels of MMP-1 (P = NS) and MMP-2 (P < .05 vs CT, 10 nmol/L, 100 nmol/L) were up-regulated after 1 mumol/L tacrolimus. Our findings confirmed that a high dose of tacrolimus does not induce interstitial collagen overexpression by gingival fibroblasts and induces up-regulation of MMPs protein levels. Interestingly, at doses corresponding to whole blood trough levels, tacrolimus did not exert any evident effect on collagen turnover pathways, suggesting that tacrolimus is likely to not affect collagen homeostasis in the gingival connective tissue compartment of FK506-immunosuppressed subjects. This effect did not seem to be dose-dependent.
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PMID:A therapeutic dose of FK506 does not affect collagen turnover pathways in healthy human gingival fibroblasts. 1858 21

Cucumber seedlings infected with cucumber mosaic virus (CMV) contain a virus-induced RNA-dependent RNA polymerase (RNA replicase) in both soluble and particulate fractions of plant extracts; no such activity is found in uninfected plants. The extensive purification and characterization of the soluble RNA replicase has been described (R. Kumarasamy and R. H. Symons, 1979, Virology 96, 622-632) while the characterization of the particulate enzyme is reported here. The particulate RNA replicase was solubilized-by incubation in 150 mM MgS04 at 37 degrees for 5 min; both magnesium and sulfate ions were required for maximum release. Only 5-7% of the protein of the particulate fraction was released under these conditions either in the presence or absence of a variety of salts. The solubilized enzyme was purified about 60-fold by a three-step procedure to a specific activity similar to that of the 10,000-fold purified soluble enzyme. CMV RNA and poly(C)-copying activities copurified through the three steps. Purification was followed by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate of the in vitro 3H-labeled proteins. The most prominent virus-induced components were two proteins of Mr 100,000 and 110,000 which correspond in size to two virus-induced proteins in the extensively purified soluble RNA replicase. The Mr 110,000 protein comigrated with the full length in vitro translation product of CMV RNA 2 while the Mr 100,000 was detectably larger than the full length in vitro translation product of CMV RNA 1.
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PMID:Cucumber mosaic virus-induced RNA replicase: solubilization and partial purification of the particulate enzyme. 1863 80

The extensively purified RNA-dependent RNA polymerase (RNA replicase) induced by infection of cucumber seedlings with cucumber mosaic virus (CMV) was investigated to determine whether the enzyme contains full-length translation products of the CMV genomal RNAs. Two experimental approaches were used. (1) RNA replicase preparations from plants infected with three strains of CMV (Q, P, and T) all had almost identical polypeptide compositions, which included a major polypeptide of relative mobility on sodium dodecyl sulfate-polyacrylamide gels of Mr, 100,000, and two other proteins, Mr 110,000 and Mr, 35,000, present in smaller, varying amounts. These polypeptides were unique to enzyme preparations from CMV-infected plants and had similar electrophoretic mobilities to the translation products of Q-CMV RNAs 1, 2, and 3, respectively. Analysis of the in vitro translation products of the corresponding RNAs of the other two strains of CMV showed, however, that those of P-CMV RNA 2 and T-CMV RNAs 1 and 3 varied significantly in size from the translation products of the corresponding Q-CMV RNAs. (2) Peptide mapping studies, using digestion with the V8 protease from Staphylococcus aureus or cleavage with CNBr, confirmed that none of these three components of the highly purified RNA replicase was a translation product of the CMV RNAs. The work reported in this paper, therefore, showed that the full-length translation products of the genomal RNAs of CMV were absent from the RNA replicase induced by this virus after solubilization and extensive purification.
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PMID:Highly purified cucumber mosaic virus-induced RNA-dependent RNA polymerase does not contain any of the full length translation products of the genomic RNAs. 1863 45

Prophenoloxidase (proPO) is a melanin-synthesising enzyme that plays important roles in immune responses by crustaceans. Previously, we cloned and characterized proPO-I from white shrimp, Litopenaeus vannamei. In the present study, a novel prophenoloxidase-II (proPO-II) cDNA was also cloned from haemocytes of L. vannamei using oligonucleotide primers and reverse-transcriptase polymerase chain reaction (RT-PCR). Both 3'- and 5'-regions were isolated by the rapid amplification of complementary (c)DNA end (RACE) method. The 2504-bp cDNA contained an open reading frame (ORF) of 2073 bp, an 84-bp 5'-untranslated region, and a 347-bp 3'-untranslated region containing the poly A tail. The molecular mass of the deduced amino acid sequence (691 amino acids) was 78.8 kDa with an estimated pI of 6.07. It contains two putative tyrosinase copper-binding motifs and a conserved C-terminal region common to all known proPOs. Comparisons of the amino acid sequences showed that white shrimp proPO-II is more closely related to the proPO of other penaeids than to that of crayfish, lobsters, crab, or a freshwater prawn, and is the ancestor type of known penaeid proPOs. proPO-I and proPO-II messenger (m)RNAs of shrimp were located on different loci, and were constitutively expressed mainly in haemocytes. The transcriptional regulation of these two proPOs in shrimp at different molt stages, those administered dietary sodium alginate, and those challenged with Vibrio alginolyticus were surveyed. The results showed that the proPOs may be directly involved in the acute-phase immune defence, and proPO-II may contribute earlier to immune defence in shrimp injected with V. alginolyticus, and it may be regulated by ecdysone. However, a similar effect was found by stimulating proPO-I and proPO-II mRNA expression in shrimp fed a sodium alginate-containing diet. Results of this study provide a basis for developing a comprehensive understanding of expression/function relationships of individual proPOs in shrimp.
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PMID:A second proPO present in white shrimp Litopenaeus vannamei and expression of the proPOs during a Vibrio alginolyticus injection, molt stage, and oral sodium alginate ingestion. 1898 57

Nitric oxide (NO) is a bioactive molecule involved in several growth and developmental processes in plants. These processes are mostly characterized by changes in primary and secondary metabolism. Here, the effect of NO on cellulose synthesis in tomato (Solanum lycopersicum) roots was studied. The phenotype of roots, cellulose content, the incorporation of 14C-glucose into cellulosic fraction and the expression of tomato cellulose synthase (CESA) transcripts in roots treated with the NO donor sodium nitroprusside (SNP) were analysed. Nitric oxide affected cellulose content in roots in a dose dependent manner. Low concentrations of SNP (pmoles of NO) increased cellulose content in roots while higher concentrations of SNP (nmoles of NO) had the opposite effect. This result correlated with assays of 14C-glucose incorporation into cellulose in roots. The effect of NO on 14C-glucose incorporation into cellulose was transient and reversible. Microscopic analysis of roots suggested that NO affected primary cell wall cellulose synthesis. Three tomato cellulose synthase (SICESA) transcripts were identified. Reverse transcriptase polymerase chain reaction experiments were carried out and indicated that SICESA1 and SICESA3 levels were affected by high NO concentrations. Together, these results support the hypothesis that variations in NO levels influence cellulose synthesis and content in roots.
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PMID:Nitric oxide: an active nitrogen molecule that modulates cellulose synthesis in tomato roots. 1863 43

The C-terminal domain protein (amino-acid residues 535-759) of the PB2 subunit of the RNA-dependent RNA polymerase from the highly pathogenic influenza A virus was expressed as a soluble protein in Escherichia coli and crystallized using sodium formate as a precipitant. Data sets were collected from crystals of native and selenomethionine-substituted protein on the KEK NW12 beamline at the Photon Factory and the crystals diffracted to a maximum resolution of 2.44 A for the SeMet-derivative crystal. The native crystals were found to belong to space group P3(2)21, with unit-cell parameters a = b = 52.5, c = 156.3 A. The Matthews value (V(M)) was 2.7 A(3) Da(-1), assuming the presence of one molecule in the asymmetric unit. The SeMet-derivative crystals were found to belong to the same space group, with unit-cell parameters a = b = 52.6, c = 156.4 A. Attempts are being made to solve the structure by multi-wavelength anomalous dispersion phasing.
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PMID:Crystallization and X-ray diffraction analysis of the RNA primer/promoter-binding domain of influenza A virus RNA-dependent RNA polymerase PB2. 1919 6

The ineffectiveness of anticancer drugs is frequently observed in cancer chemotherapy. The resistance of tumor cells to various cytotoxic drugs is defined as multidrug resistance (MDR). The purpose of our present study was to investigate the inhibitory effects of L1EPO synthesized by our group on P-glycoprotein (P-gp)-mediated MDR in K562/A02 and KBv200 cell lines, which expressed high levels of P-gp. Both the cytotoxicity of the compound and its ability to inhibit K562/A02 and KBv200 cells were determined by sulforhodamine B sodium salt (SRB) assay. Morphologic apoptosis was detected by Hoechst33342 staining assay. Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect mdr-1 gene transcription, and Western blot assay was used to assess P-gp expression. Interestingly, we found that the K562/A02 cell line was rendered resistant toward Adriamycin but not towards L1EPO when compared with the parental cells. Furthermore, L1EPO could down-regulate the mdr-1 gene, and it reduced the expression of P-gp and displayed a perfect dose dependence. Moreover, it had less cytotoxicity in normal human cell lines (fibroblast, VEC), GI(50)>10 micromol/l. Consequently, L1EPO has the potential to overcome P-glycoprotein-mediated MDR in the K562/A02 cell line.
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PMID:L1EPO, a novel podophyllotoxin derivative overcomes P-glycoprotein-mediated multidrug resistance in K562/A02 cell line. 1933 92

Gamma-hydroxybutyric acid (GHB), a drug of abuse, is a substrate of monocarboxylate transporters (MCTs). Sodium-coupled monocarboxylate transporter 1 (SMCT1; SLC5A8) is expressed in kidney, thyroid gland, neurons, and intestinal tract and exhibits substrate specificity similar to that of the proton-dependent MCT (SLC16A) family. The role of SMCT1 in GHB disposition has not been determined. In this study we characterized the driving force, transport kinetics, and inhibitors of GHB uptake, as well as expression of SMCT and MCT isoforms, in rat thyroid follicular (FRTL-5) cells. GHB, as well as the monocarboxylates butyrate and d-lactate, exhibited sodium-dependent uptake at pH 7.4, which could be described with a simple Michaelis-Menten equation plus a diffusional component [K(m) 0.68 +/- 0.30 mM, V(max) 3.50 +/- 1.58 nmol . mg(-1) . min(-1), and diffusional clearance (P) 0.25 +/- 0.08 microl . mg(-1) . min(-1)]. In the absence of sodium, GHB uptake was significantly increased at lower pH, suggesting proton-gradient dependent transport. Reverse transcriptase-polymerase chain reaction and Western analyses demonstrated the expression of SMCT1, MCT1, and MCT2 in FRTL-5 cells, supporting the activity results. Sodium-dependent GHB uptake in FRTL-5 cells was inhibited by MCT substrates (d-lactate, l-lactate, pyruvate, and butyrate), nonsteroidal anti-inflammatory drugs (ibuprofen, ketoprofen, and naproxen), and probenecid. IC(50) values for l-lactate, ibuprofen, ketoprofen, and probenecid were 101, 31.6, 64.4, and 380 muM, respectively. All four inhibitors also significantly inhibited GHB uptake in rat MCT1 gene-transfected MDA/MB231 cells, suggesting they are not specific for SMCT1. Luteolin and alpha-cyano-4-hydroxycinnimate represent specific proton-dependent MCT inhibitors. Our findings indicate that GHB is a substrate for both sodium- and proton-dependent MCTs and identified specific inhibitors of MCTs.
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PMID:The drug of abuse gamma-hydroxybutyrate is a substrate for sodium-coupled monocarboxylate transporter (SMCT) 1 (SLC5A8): characterization of SMCT-mediated uptake and inhibition. 1938 57

Several studies indicate an essential role of the heterodimer Tas1R1-Tas1R3 for monosodium l-glutamate (MSG) detection, although others suggest alternative receptors. Human subjects show different taste sensitivities to MSG, and some are unable to detect the presence of glutamate. Our objective was to study possible relations between phenotype (sensitivity to glutamate) and genotype (polymorphisms in candidate glutamate taste receptors tas1r1, tas1r3, mGluR4, and mGluR1) at the individual level. The sensitivity was measured with a battery of tests to distinguish the effect of sodium ions from the effect of glutamate ions in MSG. A total of 142 genetically unrelated white French subjects were categorized into 27 nontasters (specific ageusia), 21 hypotasters, and 94 tasters. Reverse transcriptase polymerase chain reaction and immunohistochemistry showed expression of tas1r1, tas1r3, and alpha-gustducin in fungiform papillae in all 12 subjects tested, including subjects who presented specific ageusia for glutamate. Amplification and sequencing of cDNA and genomic DNA allowed the identification of 10 nonsynonymous single nucleotide polymorphisms (nsSNPs) in tas1r1 (n = 3), tas1r3 (n = 3), and mGluR1 (n = 4). In our sample of subjects, the frequencies of 2 nsSNPs, C329T in tas1r1 and C2269T in tas1r3, were significantly higher in nontasters than expected, whereas G1114A in tas1r1 was more frequent in tasters. These nsSNPs along with minor variants and other nsSNPs in mGluR1, including T2977C, account for only part of the interindividual variance, which indicates that other factors, possibly including additional receptors, contribute to glutamate sensitivity.
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PMID:Nonsynonymous single nucleotide polymorphisms in human tas1r1, tas1r3, and mGluR1 and individual taste sensitivity to glutamate. 1957 Dec 23


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