Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.6 (RNA polymerase)
 34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An RNA-dependent RNA polymerase activity has been demonstrated for spring viremia of carp virus (SVCV). The optimal temperature for in vitro synthesis of RNA was 20 to 25 degrees C. The SVCV enzyme activity was stimulated when the methyl donor S-adenosyl-L-methionine was included in the reaction mixture. S-adenosyl-L-methionine was not particularly effective in stimulating the virion RNA polymerase activity of vesicular stomatitis virus or pike fry rhabdovirus. The 5' nucleotide of the SVCV viral RNA is pppAp.
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PMID:Spring viremia of carp virus RNA and virion-associated transcriptase activity. 56 17

S-Adenosyl-L-methionine (SAM), a methyl donor, and its analogue S-adenyl-L-homocysteine (SAH), an inhibitor of methylation, stimulate the activity of spring viraemia of carp virus (SVCV) virion transcriptase. The stimulation observed for SVCV is analogous to that observed previously (Furuichi, 1974, 1978) for a totally unrelated virus, cytoplasmic polyhedrosis virus (CPV). In the absence of exogenous SAM, RNA with 5'-methylated termini (presumptive GpppAmpAp) was produced, indicating that SVCV has an endogenous methyl donor. Significantly less methylated termini were produced when SVCV nucleocapsids were used to prime in vitro transcription reactions, suggesting that the majority of the endogenous methyl donor is not associated with the nucleocapsid. Partial removal of endogenous methyl donor by preparing nucleocapsids did not have any effect on the degree of stimulation by exogenous SAM or SAH. We conclude from this study that SAH has two effects on SVCV transcription, inhibition of methylation and stimulation of transcription.
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PMID:Stimulation of transcription by S-adenosyl-L-homocysteine and virion-encapsidated methyl donor in spring viraemia of carp virus. 727 15

During seasonal acclimatization of eurythermal fish, the nucleolus of the hepatocyte undergoes ultrastructural reprogramming. In winter acclimatized carp, the nucleolar components are segregated, a condition that suggests a decreased transcription of rRNA. The same nucleolar reorganization was observed when pituitary cells from winter- and summer-acclimatized carp were examined. In situ analyses of nucleolar RNA revealed a marked lowering of RNA content in the segregated nucleoli. Accordingly, in vitro synthesis of RNA was shown to be significantly lower in pituitary tissue from cold-acclimatized fish where precursor accumulated. Conversely, in pituitary tissue from summer-adapted fish the rate and extent of synthesis and of rRNA processing was notably higher. The involvement of pre-rRNA processing events during seasonal acclimatization was corroborated by the strong differences of U3 RNA content detected by in situ hybridization in pituitary cells from summer- and winter-fish. When RNA polymerase I activity from both acclimatized states were assayed, no differences were detected. Thus, it appears that in fish RNA polymerase I itself does not play an important role in the control of nucleolar gene expression and the nucleolar gene expression reprogramming that the seasonal rearrangement represents might involve, among the many nucleolar-specific proteins, transcription factors.
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PMID:Reprogramming of nucleolar gene expression during the acclimatization of the carp. 751 43

A human autoimmune serum to nucleolus organizer regions (NORs) has been used to localize these structures at the light microscopic level in carp and trout tissue culture cells. In interphase cells, the immunofluorescence pattern indicates that the NORs autoantigens are contained exclusively within the nucleolus of carp epithelial (EPC) and trout gonad (RTG) cells. This fluorescence is punctuate rather than uniform, and presumably represents transcriptional complexes of ribosomal DNA. During mitosis, the autoantigens are detected by immunofluorescence microscopy at the chromosomal nucleolus organizer regions of condensed chromosomes, indicating that a considerable quantity of the molecule(s) remains bound to the ribosomal RNA genes. The major nucleolus autoantigen, defined in mammals as the upstream ribosomal binding factor (UBF), has been identified on immunoblots as a 90 kDa protein in extracts from fish cell lines and tissues. Thus, NORs appear to function as nucleation centers for ribosomal RNA together with a complex set of well-conserved protein factors, such as UBF. Our results suggest evolutionary conservation from fish to mammals with respect to ribosomal RNA biosynthesis driven by RNA polymerase I.
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PMID:Immunodetection of the ribosomal transcription factor UBF at the nucleolus organizer regions of fish cells. 795 74

Connexin26 (Cx26) is a member of the family of integral membrane proteins that normally form intercellular gap junctional channels. We have used Western blotting, immunofluorescence, immunoelectron microscopy, and single-cell reverse-transcriptase polymerase chain reaction amplification (RT-PCR) to analyze the expression and cellular localization of Cx26 in the carp retina. In the outer plexiform layer, strong clustered Cx26 immunolabeling was concentrated at and restricted to the terminal dendrites of horizontal cells. Single-cell RT-PCR confirmed the expression of Cx26 in carp retinal horizontal cells. 248-bp fragments amplified from cDNAs of four different horizontal cells were cloned and each nucleotide sequence encodes a protein fragment (AA 104-185) with highly significant homology to rat and mouse Cx26. Immunoelectron microscopy revealed that only the invaginating dendrites of horizontal cells in intimate lateral association with the presynaptic ribbon complex were labeled. No labeling was found at the photoreceptor membrane and there was no septalaminar structure, indicative of gap junctions, between photoreceptors and horizontal cells. The focal location of Cx26 at the membrane of the dendritic tips of horizontal cells and the lack of gap junctional morphology suggests that Cx26 might form hemichannels.
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PMID:Identification and localization of connexin26 within the photoreceptor-horizontal cell synaptic complex. 1141 91

We investigated a potential mechanism for the estrogenic properties of three chloro-s-triazine herbicides and six metabolites in vitro in several cell systems. We determined effects on human aromatase (CYP19), the enzyme that converts androgens to estrogens, in H295R (adrenocortical carcinoma), JEG-3 (placental choriocarcinoma), and MCF-7 (breast cancer) cells; we determined effects on estrogen receptor-mediated induction of vitellogenin in primary hepatocyte cultures of adult male carp (Cyprinus carpio). In addition to atrazine, simazine, and propazine, two metabolites--atrazine-desethyl and atrazine-desisopropyl--induced aromatase activity in H295R cells concentration-dependently (0.3-30 microM) and with potencies similar to those of the parent triazines. After a 24-hr exposure to 30 microM of the triazines, an apparent maximum induction of about 2- to 2.5-fold was achieved. The induction responses were confirmed by similar increases in CYP19 mRNA levels, determined by reverse-transcriptase polymerase chain reaction. In JEG-3 cells, where basal aromatase expression is about 15-fold greater than in H295R cells, the induction responses were similar but less pronounced; aromatase expression in MCF-7 cells was neither detectable nor inducible under our culture conditions. The fully dealkylated metabolite atrazine-desethyl-desisopropyl and the three hydroxylated metabolites (2-OH-atrazine-desethyl, -desisopropyl, and -desethyl-desisopropyl) did not induce aromatase activity. None of the triazine herbicides nor their metabolites induced vitellogenin production in male carp hepatocytes; nor did they antagonize the induction of vitellogenin by 100 nM (EC(50) 17beta-estradiol. These findings together with other reports indicate that the estrogenic effects associated with the triazine herbicides in vivo are not estrogen receptor-mediated, but may be explained partly by their ability to induce aromatase in vitro.
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PMID:Effects of chloro-s-triazine herbicides and metabolites on aromatase activity in various human cell lines and on vitellogenin production in male carp hepatocytes. 1167 67

The complete genome of spring viremia of carp virus (SVCV) was cloned and the sequence of 11019 nucleotides was determined. It contains five open reading frames (ORF's) encoding for the nucleoprotein N; phosphoprotein P; matrix protein M; glycoprotein G; and the viral RNA dependent RNA polymerase L. Genes are organised in the order typical for rhabdoviruses: 3'-N-P-M-G-L-5'. The short leader and trailer regions of SVCV exhibit inverse complementarity and are similar to the respective 3' and 5' ends of the genome of vesicular stomatitis virus. To verify the predicted open reading frames proteins were expressed in bacteria and analysed with a polyclonal anti-SVCV serum. Furthermore, monospecific antisera against the distinct viral proteins were generated. Comparison of genome and protein confirm the assignment of SVCV to the genus Vesiculovirus.
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PMID:Determination of the complete genomic sequence and analysis of the gene products of the virus of Spring Viremia of Carp, a fish rhabdovirus. 1190 Aug 42

This study demonstrated that a galactose-binding protein (GBP) produced by a fish pathogenic water mold, Aphanomyces piscicida, activates carp leukocytes. Leukocytes were separated from the head kidney and peripheral blood using Percoll density centrifugation. A flow cytometric analysis revealed that GBP binds with many cells and a variety of cell types including lymphocytes, granulocytes and thrombocytes. Intracellular calcium flux of the peripheral blood leukocytes induced by stimulation with GBP was confirmed by counting the fluo-3 loaded cells whose fluorescence increased after the stimulation using flow cytometry. The percentage of cells in which a calcium flux was induced peaked 1 min after the stimulation. Approximately 6% of the cells specifically responded 1 min after the stimulation. The proliferation response was determined by the level of BrdU uptake by the leukocytes after the stimulation. Cell proliferation was observed 2, 4 and 6 days after stimulation with GBP. The expression of cytokines IL-1beta and TGF-beta1 in the peripheral blood leukocytes, after the stimulation was evaluated by a semi-quantitative reverse-transcriptase polymerase chain reaction. Increased expression of IL-1beta was observed 4h after stimulation with GBP. Variation of TGF-beta1 expression under the same conditions was not observed. The kinetics of intracellular calcium flux and the level of IL-1beta expression induced by GBP stimulation were different from those induced by phytohemagglutinin stimulation. These results confirmed that GBP is a pathogenic microbial component that can induce cell activation. GBP seems to induce the inflammatory response observed in the Aphanomyces infection.
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PMID:Activation of carp leukocytes by a galactose-binding protein from Aphanomyces piscicida. 1190 25

A complementary DNA (cDNA) encoding the eggshell zona radiata protein (RBTZR: AF407574) has been cloned from the liver of estradiol-17beta (E(2))-treated rainbow trout (Oncorhynchus mykiss) by reverse-transcriptase polymerase chain reaction (RT-PCR). A set of degenerate primers homologous to the highly conserved cysteine-rich region of the zona radiata protein gene from salmon, winter flounder, medaka and carp were used for the initial RT-PCR. The resulting PCR product was cloned, sequenced and identified as the Zrp gene fragment based on amino acid sequence similarities. Based on the Zrp sequence from the initial PCR, a pair of gene-sequence primers was designed for 3'- and 5'- random amplification of cDNA ends (RACE). Cloning and sequencing of RACE products showed a 1349-bp Zrp gene encoding a 403-amino acid protein with a theoretical molecular mass of approximately 45 kDa. Alignment of the deduced amino acid sequence reveals that RbtZR is similar to piscine and mammalian zona pellucida proteins. The RbtZR gene, together with the estrogen receptor (ER) and vitellogenin (Vtg) genes, was further characterized and comparatively studied for transcriptional and translational expression in xenoestrogen- (nonylphenol, NP) and E(2)-treated juvenile rainbow trout in a time-course experiment. Northern and slot blot analysis showed that the RbtZR mRNA was expressed, in parallel with the ER and Vtg mRNA, in both NP- and E(2)-treated juvenile rainbow trout. Indirect enzyme-linked immunosorbent assay (ELISA) using monoclonal antibody raised against Atlantic salmon Zrp indicated the translational expression of RbtZR protein in blood plasma samples from NP- and E(2)-treated juvenile trout. The differential time-dependent transcriptional and translational expression and use of Zrp, ER and Vtg as sensitive biomarkers in environmental monitoring of endocrine disrupters in fish is discussed.
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PMID:Molecular cloning of rainbow trout (Oncorhynchus mykiss) eggshell zona radiata protein complementary DNA: mRNA expression in 17beta-estradiol- and nonylphenol-treated fish. 1203 56

The presence of the female-specific yolk protein precursor vitellogenin in blood and liver from male fish is widely used as an indicator of endocrine disruption. We studied the induction of vitellogenin mRNA in liver from several species of fish, both maintained in fish tanks or captured in the wild. Our procedure requires minute amounts of liver samples (down to 50 mg), and can be applied to field samples if the appropriate RNA-stabilisation agent is used. We used reverse-transcriptase PCR and quantitative real-time PCR for detection and precise quantitation of vitellogenin mRNA levels. Male mummichog (Fundulus heteroclitus) exposed to 17 beta-estradiol contained levels of vitellogenin mRNA up to 30 times higher than in untreated females and treatment with nonylphenol resulted in a weak but consistent induction of this transcript. We also studied levels of vitellogenin mRNA in a population of common carp (Cyprinus carpio) from the Anoia river, a river known for its high levels of estrogenic alkylphenols. The results were consistent with recorded data for fish from this sampling site. Finally, we also detected vitellogenin mRNA in barbs (Barbus graellsi), a species for which no vitellogenin sequence was available. The use of mRNA quantitation techniques for analysis of feral and cultured fish of different species opens the possibility of more precise detection and further control of the noxious effects of contaminants on the local fauna exposed to them.
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PMID:Use of vitellogenin mRNA as a biomarker for endocrine disruption in feral and cultured fish. 1461 90


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