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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have cloned the cDNA encoding the voltage-dependent K+ channel Kv2.1 from human brain (hKv2.1). RNase protection and RT-PCR (reverse transcriptase-PCR) experiments reveal abundant Kv2.1 transcripts in human brain with virtually no expression detectable in human heart. hKv2.1 has been stably transfected into a human glioblastoma cell line, and transformed cells display large, slowly activating outward currents. The kinetics, steady-state activation and inactivation parameters, and external tetraethylammonium sensitivity were all similar to those described previously for hKv2.1 channels transiently expressed in Xenopus oocytes or other mammalian cell lines. A number of dopamine receptor antagonist/antipsychotic agents were shown to block hKv2.1. Trifluoperizine, trifluperidol and pimozide produced time-dependent blockade of hKv2.1 with IC50 values of approx. 1-2 microM. The diphenylbutylpiperidine fluspirilene was shown to be 4-5-fold more potent than the other agents tested inhibiting hKv2.1 current with an IC50 value of 297 nM. The block produced by fluspirilene was both time- and frequency-dependent. Furthermore, fluspirilene (1 microM) shifted the midpotential of the hKv2.1 steady-state inactivation curve by approx. 15 mV in the hyperpolarizing direction. These results demonstrate the usefulness of this transfection system for the pharmacological characterization of hKv2. 1. Fluspirilene proved to be a relatively potent blocker of hKv2.1 and may provide a useful starting point for the development of more potent and selective agents active against this brain K+ channel.
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PMID:Stable expression and characterization of the human brain potassium channel Kv2.1: blockade by antipsychotic agents. 924 64

Latent membrane protein 1 (LMP 1) is one of two Epstein-Barr virus (EBV)-encoded proteins that expressed in nasopharyngeal carcinoma (NPC) cells. Previous studies showed that a 3.5-kb transcript of the LMP 1 gene, in addition to the 2.8-kb transcript, was detected in a B95-8-EBV-containing, nude mice-passaged NPC tumor, C15. This indicated that a transcript was initiated from a region 5' to the putative promoter, ED-L1. We have isolated an EBV variant from a NPC tissue, and this virus strain contained a more pathogenic LMP 1 gene. DNA sequence analysis of the 5'-upstream region showed distinct variations as compared to that of B95-8 strain. To test if the LMP 1 gene of the NPC strain also contained an upstream promoter, we generated a series of deletion plasmids encompassing positions -1,030 to +20 of the LMP 1 promoter and tested for their abilities to drive the expression of the reporter gene in human epithelial cell lines, C-33A and NPC-TW076. We found that the region between -643 and -496 contained a promoter activity that was approximately five-fold higher than the putative promoter, ED-L1. This region between -643 and -496 was designated as ED-L1E. C-33A cells containing the genomic clone pT7(E) or the clone that had deleted a 94-bp ED-L1 sequence (delta94) was used to determine the transcription initiation sites by RNase protection assay. Results showed that a transcription initiation site was located at nucleotide 170,099 ("A") of EBV genome. The transcript was expressed in NPC biopsies and in human primary normal epithelial cells transfected with pT7(E) and delta94, respectively, as examined by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. Furthermore, the ED-L1E was not regulated by the EBV-encoded nuclear antigen 1-mediated transcriptional enhancer family of repeats (FR) in C-33A cells. Our results suggested that the ED-L1E was specifically activated in epithelial cells. The biological significance of the selective usage of the ED-L1E promoter was discussed.
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PMID:Identification of a promoter for the latent membrane protein 1 gene of Epstein-Barr virus that is specifically activated in human epithelial cells. 926 Sep 26

Retroviral RNases H are similar in sequence and structure to Escherichia coli RNase HI and yet have differences in substrate specificities, metal ion requirements, and specific activities. Separation of reverse transcriptase (RT) into polymerase and RNase H domains yields an active RNase H from murine leukemia virus (MuLV) but an inactive human immunodeficiency virus (HIV) RNase H. The "handle region" present in E. coli RNase HI but absent in HIV RNase H contributes to the binding to its substrate and when inserted into HIV RNase H results in an active enzyme retaining some degree of specificity. Here, we show MuLV protein containing the C-terminal 175 amino acids with its own handle region or that of E. coli RNase HI has the same specific activity as the RNase H of RT, retains a preference for Mn2+ as the cation required for activity, and has association rate (KA) 10% that of E. coli RNase HI. However, with model substrates, specificities for removal of the tRNAPro primer and polypurine tract stability are lost, indicating specificity of RNase H of MuLV requires the remainder of the RT. Differences in KA, while significant, appear insufficient to account for the differences in specific activities of the bacterial and viral RNases H.
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PMID:The isolated RNase H domain of murine leukemia virus reverse transcriptase. Retention of activity with concomitant loss of specificity. 926 41

We established a differentiation-inducible preadipocyte cell line, designated A54 preadipocytes, from C3H10T1/2 (10T1/2) mouse embryo fibroblasts. A54 preadipocytes had marked hematopoiesis-supporting ability in vitro but this ability was lost after terminal differentiation to adipocytes. In this study, to identify molecules that contribute to the hematopoiesis-supporting ability of A54 preadipocytes, we screened genes that were differentially expressed in A54 preadipocytes and isolated seven novel genes by reverse transcriptase polymerase chain reaction mRNA differential display. An RNase protection assay confirmed that one of these genes was expressed at high levels in parent 10T1/2 cells and A54 preadipocytes but to a much lesser extent in fully differentiated A54 adipocytes. This gene was defined as a gene that was downregulated during adipocyte differentiation-1 (drad-1). The size of drad-1 mRNA was 8.2 kb, and the gene was expressed in other mouse preadipocytes, namely, ST2 and PA6 cells, that have hematopoiesis-supporting ability. Moreover, drad-1 was also found to be expressed in bone marrow in vivo. The function of the protein encoded by drad-1 is unknown, but the expression of the gene may be useful as a molecular marker of adipocyte differentiation.
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PMID:A novel gene (drad-1) expressed in hematopoiesis-supporting stromal cell lines, ST2, PA6 and A54 preadipocytes: use of mRNA differential display. 932 94

We have examined whether alterations in the growth hormone/insulin-like growth factor-1 axis play a role in the pathogenesis of psoriasis. Serum, urine, full skin biopsies, and suction blister roofs were obtained from patients with psoriasis and from healthy controls. Serum concentrations of insulin-like growth factor-1 and insulin-like growth factor binding protein-3 were measured by radioimmunoassay. Growth hormone-binding protein was measured by ligand-mediated immunofunctional assay. Growth hormone concentration in urine was measured by an immunometric assay, and growth hormone receptor-gene expression was measured by RNase protection assay or by quantitative reverse transcriptase polymerase chain reaction in total RNA isolated from epidermal suction blister roofs. Serum concentrations of insulin-like growth factor-1 (249 +/- 12 micrograms per liter, mean +/- SEM, n = 42, and 277 +/- 21 micrograms per liter, n = 9, for psoriatic patients and controls, respectively), insulin-like growth factor binding protein-3 (3.1 +/- 0.08 mg per liter, n = 42, and 3.3 +/- 0.22 mg per liter, n = 9), growth hormone-binding protein (344 +/- 65 pmol per liter, n = 10, and 311 +/- 83 pmol per liter, n = 9), urinary growth hormone excretion during 24 h (12.8 +/- 2.7 microIU per 24 h, n = 12, and 12.3 +/- 1.6 microIU per 24 h, n = 9), and epidermal growth hormone receptor gene expression [32 +/- 12 x 10(3) mRNA transcripts per microgram total RNA (involved skin), n = 11, and 47 +/- 14 x 10(3) mRNA transcripts per microgram total RNA, n = 9] were similar in patients and controls. For insulin-like growth factor-1 and insulin-like growth factor binding protein-3 the values in psoriatic patients were also similar to those in larger control groups, n = 195 and n = 400, respectively. In addition, we found no evidence of local expression of growth hormone or prolactin in full skin punch biopsies from psoriatic involved skin by reverse transcriptase polymerase chain reaction. In conclusion, our results suggest that alterations in the growth hormone/ insulin-like growth factor-1 axis do not play a major role in the pathogenesis of psoriasis.
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PMID:No evidence for involvement of the growth hormone/insulin-like growth factor-1 axis in psoriasis. 934 96

We have cloned and functionally characterized the RNase H1 gene from D. melanogaster. The longest open reading frame consists of 5 exons that encode a 333 amino acid protein with a molecular mass of 37.1 kDa. This is the first demonstration of specific nuclease activity of a cloned RNase gene from a multicellular higher eukaryote. No additional proteins or cofactors are required for this nuclease activity. Comparison of Drosophila RNase H1 amino acid sequence to that of other cellular eukaryotic homologs reveals the presence of three evolutionarily distinct domains. The N- and C-terminal conserved domains are connected by a highly variable domain. The C-terminal domain has high amino acid similarity to bacterial RNase HI and the RNase H domain of retroviral reverse transcriptase, while the N-terminus, of unknown function, is similar to the P6 translational activator of caulimoviruses.
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PMID:Functional characterization of RNase H1 from Drosophila melanogaster. 939 56

This report describes cloning of the bovine alpha 2D-adrenergic receptor (alpha 2D-AR) gene and determination of the transcription start site, unequivocal presence of the alpha 2D-AR transcript in the retina, and pharmacological characteristics of the encoded product. Furthermore, expression of the gene in selected bovine tissues has also been scrutinized. A genomic clone was isolated from lambda EMBL3 library and a 3 kb fragment was subcloned and sequenced. This fragment contained the putative TATA box and the coding region. The encoded receptor was transiently expressed in COS cells. The recombinant receptor expressed pharmacological characteristics almost identical to the wild-type bovine retinal receptor, which were typical of the alpha 2D-AR subtype. RNase protection analysis confirmed the expression of the gene in the retina. The bovine receptor was structurally close to its rat analogue which also encodes the alpha 2D-AR, but, the highest homology was observed with the porcine receptor expressing alpha 2A-AR pharmacological characteristics. Certain structural features of the bovine gene were unique to itself and not shared by any other alpha2-AR subtype. Among the tissues tested using reverse transcriptase-polymerase chain reaction (RT-PCR), the alpha 2D-AR message was the most abundant in retina, followed by the brain and olfactory lobe. Thus, the availability of the bovine receptor gene probe will become an important additional tool in the elucidation of molecular mechanisms behind the alpha 2D-AR physiology in neurosensory processes such as those occurring in the eye and the brain.
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PMID:The bovine alpha 2D-adrenergic receptor gene: structure, expression in retina, and pharmacological characterization of the encoded receptor. 945 Jun 52

The existence of retroviral reverse transcriptases as monomers or dimers is rather intriguing. A classical example of the former is murine leukemia virus reverse transcriptase (MuLV RT), while human immunodeficiency virus type 1 (HIV-1) RT represents the latter. A careful scrutiny of the amino acid sequence alignment of the two enzymes pinpoints the region tentatively responsible for this phenomenon. We report here the construction of a chimeric enzyme containing the first 425 amino acid residues from the N-terminal domain of HIV-1 RT and 200 amino acid residues from the C-terminal domain of MuLV RT. The chimeric enzyme exists as a monomer with intact DNA polymerase and RNase-H functions.
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PMID:An enzymatically active chimeric HIV-1 reverse transcriptase (RT) with the RNase-H domain of murine leukemia virus RT exists as a monomer. 954 16

The Rel family of transcriptional activators form a large diverse group of proteins that are involved in the activation of genes involved in immunity, development, apoptosis and cancer. So far, none of the rel genes cloned in mammals appear to be required for embryonic development. We have cloned and characterized a cDNA from an embryonic cDNA library that encodes a novel Xenopus Rel protein, called Xrel3. Xrel3 is a member of the cRel subfamily and is most closely related to but distinct from other Xenopus Rel members. The expression of Xrel3 mRNA was investigated using Northern analysis, RNase protection assay, reverse transcriptase-linked polymerase chain reaction and in situ hybridization. Messages are present maternally and are slightly enriched in the equatorial region of the blastula stage embryo. At gastrulation, the accumulation of Xrel3 messages declines to undetectable levels but then increases after neurulation. In situ RNA hybridization was used to determine the spatial location of Xrel3 messenger RNA in embryos. Messages are localized to the developing forebrain, dorsal mid-hindbrain region, the inner ear primordium, or otocyst, and in the notochord. Overexpression by microinjection of Xrel3 RNA induced tumors in the developing embryo that appeared after gastrulation. The location of the tumors depended on the location of the injection site. These results suggest that Xrel3 might have a generalized role in regulation of cell differentiation in the embryo.
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PMID:Overexpression of a novel Xenopus rel mRNA gene induces tumors in early embryos. 959 16

Neuropathology is one approach to the effort to elucidate the pathophysiology of suicide. Initial neurochemical studies focusing on the roles of serotonin (5-HT) and noradrenaline (NE) abnormalities in brains of suicide victims have been somewhat inconsistent. More recently developed methodologies, including quantitative receptor autoradiography, immunoblotting, immunohistochemistry, cell morphometry, in situ hybridization, Northern analysis, solution hybridization/RNase protection assay, reverse transcriptase polymerase chain reaction, and genotyping, which have already been applied successfully in studies of other disorders of brain structure or function, are now increasingly being adopted for postmortem studies of suicide. These new strategies are adding convergent evidence for brain 5-HT and NE dysfunction in the etiology of suicide susceptibility, refining the neuroanatomical localization of this dysfunction, and in addition, implicating heretofore unsuspected candidate neurotransmitter systems in the neuropathological substrates of suicide susceptibility. It is argued here that the confluence of the availability of suitable postmortem samples and this augmentation of our armamentarium of techniques promises the attainment of important new insights into the biological underpinnings of suicide from postmortem research. It is to be hoped that this new knowledge might inspire novel pharmacotherapeutic strategies for the prevention of suicide.
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PMID:Neuropathology of suicide. A review and an approach. 961


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