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)

Complementary DNA (cDNA)-oligodeoxythmidylate-celluloses were prepared from cDNA copies of polysomal messenger RNA (mRNA) of Novikoff hepatoma, normal rat liver, and regenerating rat liver. cDNA synthesis with reverse transcriptase was approximately 46% with respect to input mRNA with oligodeoxythymidylate-cellulose primer. The cDNA's of normal liver, regenerating liver, and Novikoff hepatomas were used as affinity matrices for hybridization of different mRNA species. Under the conditions used, degradation of mRNA was not detected. After normalization for homologous hybridization efficiency, 53 and 65% of the Novikoff hepatoma mRNA bound to normal liver and regenerating liver cDNA's. Under these conditions an average of 82% of mRNA of normal liver bound to regenerating liver cDNA, and 92% of regenerating liver mRNA bound to normal liver cDNA. The bound and unbound mRNA's were analyzed by translation in the wheat germ system; 2-D gel analysis of the proteins synthesized in the wheat germ system indicated that the cDNA affinity columns selectively adsorbed some mRNA species.
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PMID:Adsorption of messenger RNA of Novikoff hepatoma, normal liver, and regenerating liver on complementary DNA-cellulose affinity matrices. 7 Dec

Full-length radiolabeled albumin and alpha-fetoprotein (AFP) cDNAs were synthesized from pure albumin and AMP mRNA preparations by using avian myeloblastosis virus reverse transcriptase (RNA-dependent DNA polymerase). The cDNAs have been used to quantitate the number of albumin and AFP genes in different rat tissues by two independent methods, both of which yielded similar results. First, the kinetics of the association of these cDNAs with nuclear DNA from rat liver, rat kidney, and Morris hepatoma 7777 under conditions of vast DNA excess indicated that the albumin and AFP mRNA's are transcribed from "nonrepetitive DNA." Second, saturation hybridization experiments in which a constant amount of rat liver DNA or Morris hepatoma 7777 was hybridized with increasing amounts of cDNA to albumin mRNA have shown the presence of 1--2 albumin genes per rat haploid genome. The number of AFP genes obtained in similar titration experiments was approximately 2--3. This was true whether rat liver DNA or hepatoma 7777 DNA was used in the reassociation experiments. When high molecular weight DNA preparations from both these tissues were digested with the restriction endonuclease EcoRI and the fragments were transferred to a nitrocellulose filter, the albumin and AFP [32P]cDNA probes hybridized to different sets of DNA fragments. However, each probe gave the same hybridization pattern whether Buffalo rat liver DNA or hepatoma 7777 DNA was utilized.
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PMID:alpha-Fetoprotein and albumin genes of rats: no evidence for amplification-deletion or rearrangement in rat liver carcinogenesis. 8 3

Rat alpha 1-fetoprotein mRNA was isolated and purified to apparent homogeneity by means of immunoadsorption and oligo (dT) cellulose affinity chromatography. Purified AFP mRNA migrated as a 21S peak in 2.5% SDS-polyacrylamide gels. The translation product of this mRNA in micrococcal nuclease treated reticulocyte lysate was identified as AFP by specific immunoprecipitation, SDS-gel electrophoresis and tryptic digestion analysis. DNA complimentary to AFP mRNA was synthesized with avian meyloblastosis virus RNA-dependent DNA polymerase. This AFP cDNA was used as a probe to quantitate AFP mRNA in the developing rat liver and to compare the complexity and diversity of AFP mRNA derived from the normal rat liver and Morris hepatoma 7777. We found that the amount of functional AFP mRNA is decreasing during liver development. There is very little, if any, AFP mRNA in the adult rat liver. A high degree of homology between the AFP mRNA sequences of yolk sac and hepatoma was also found.
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PMID:alpha 1-Fetoprotein mRNA of rat yolk sac and hepatoma. 9 Nov 59

Electron microscopic examination of hepatoma tissue culture (HTC) cells revealed low numbers of intracisternal type A particles (IAP) and type C viruses. Exposure of HTC cells to either 10(-4) or 10(-5) M5-bromo-2'-deoxyuridine (BUdR) caused a more than 50-fold increase in the number of IAP observed with the electron microscope. The number of IAP increased after only 2 days of growth in 10(-5) M BUdR, whereas 3 days of growth in 10(-4) m BUdR were necessary to observe an increase. A 2-day pulse of 10(-4) M BUdR was also sufficient to cause the increase in type A particles, provided the cells were continued in culture for another 2 days. Unlike many other cell lines, HTC cells treated with BUdR did not show an increase in type C viruses. This conclusion was based on the observations that BUdR treatment caused no detectable increase in extracellular particulate viral reverse transcriptase or in viral RNA complementary to a DNA probe made to a rat endogenous type C virus (RaLV).
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PMID:Induction of intracisternal type A particles by 5-bromo-2'-deoxyuridine in rat hepatoma cells. 20 99

Hepatitis B virus (HBV) contains a particle-associated DNA polymerase/reverse transcriptase activity encoded by the P (pol) open reading frame. Due to its low abundance, the corresponding protein has so far escaped direct detection and structural analysis. As a first step to overcome these difficulties, a series of recombinant vaccinia viruses was constructed and used for the synthesis in human hepatoma cells of both the authentic full length protein and of its functional domains. Pulse chase experiments demonstrated that the P-proteins had very short half lives in striking contrast to the viral core protein expressed in parallel with the same system. No evidence was obtained for a specific proteolytic processing of the P-protein as occurring with retroviral pol gene products. Overexpression of P-protein by recombinant vaccinia viruses was then employed to develop a highly sensitive detection method based on the in vitro phosphorylation of newly introduced target sites for protein kinase A. The usefulness of this method was demonstrated in the analysis of encapsidated P-gene products that were transiently expressed from an appropriately modified HBV genome. The results obtained indicate that the P-protein acts unprocessed, at least during the initial steps of nucleocapsid assembly and reverse transcription, and that a fraction of the P-protein molecules is linked as such to the viral DNA. Direct detection of the hepadnaviral P-protein by in vitro phosphorylation should greatly facilitate future analyses on P-protein structure and function.
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PMID:Expression of the P-protein of the human hepatitis B virus in a vaccinia virus system and detection of the nucleocapsid-associated P-gene product by radiolabelling at newly introduced phosphorylation sites. 137 44

We investigated the presence of the hepatitis C virus (HCV) genome in liver tissues of eight different patients with hepatocellular carcinoma by using the reverse transcriptase polymerase chain reaction (PCR) method. RNA was extracted separately from cancerous and peripheral noncancerous portions of the liver tissues of each patient. For reverse transcriptase PCR, we used sets of primers derived either from nonstructural region 3 (the NS3 region) or from the nucleocapsid-envelope (C/E) region of the HCV genome. The nucleotide sequences of the amplimers were directly determined without subcloning. Of 16 samples tested, cDNA of the HCV genome was detected in 2 cancerous tissues and in 4 noncancerous tissues by either pair of primers. Nucleotide sequences of HCV cDNA fragments amplified from cancerous and peripheral noncancerous tissues from the same patients were identical. However, 4.4 to 6.3% and 7.5 to 11.3% sequence variation was observed in NS3 and C/E regions, respectively, among cDNA fragments from different patients. The result indicated that the HCV genome detected in a given patient is distinguishable from that in others by a simple direct nucleotide sequencing of the reverse transcriptase PCR products.
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PMID:Discrimination of hepatitis C virus in liver tissues from different patients with hepatocellular carcinomas by direct nucleotide sequencing of amplified cDNA of the viral genome. 166 47

Hepatitis B virus (HBV) is the causative agent of hepatocellular carcinoma (HCC) in man. The HBV genome is a circular partially double-stranded DNA molecule of about 3.2 kb. The HBV genome contains four structural genes coding for the HBV envelope (HBsAg) and core (HBcAg/HBeAg) proteins, endogenous DNA-polymerase with the additional enzymatic activity of a reverse transcriptase and polypeptide X functioning as a trans-activator of cellular and viral genes. HBV DNA integration in the genomes of HCCs and hepatocytes of HBV carriers is an important evidence establishing a relationship between the HBV infection and the development of HCC. The mechanism of HBV DNA integration into the cellular genome and the possible role of integrated HBV DNA sequences in the malignant transformation of hepatocytes are discussed.
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PMID:[Human hepatitis B virus and hepatocellular carcinoma]. 215 10

Duck hepatitis B virus (DHBV) is produced in small amounts following transfection of human hepatoma or hepatoblastoma cell lines with cloned viral DNA. In a search for better hosts for DHBV replication, two avian liver cell lines were investigated. One of these cell lines, LMH, produced 5 to 10 times more DNA replicative intermediates and 10 to 20 times more infectious DHBV than did either of the two human cell lines, HuH-7 and Hep G2. Utilization of cell lines in genetic analyses of virus replication is often dependent upon obtaining efficient complementation between cotransfected viral genomes. We assayed transcomplementation of a viral polymerase (pol) gene mutant, which is rather inefficient in transfected human cells, and found that viral DNA synthesis was at least 20 times more efficient following cotransfection of LMH cells than in similarly transfected HuH-7 cells. Recombination, a potential interpretation problem in complementation assays, occurred at low levels in the cotransfected cultures but was substantially reduced or eliminated by creation of an LMH subline stably expressing the viral polymerase. This cell line, pol-7, supported the replication of DHBV pol mutants at ca. 10 to 15% of the level of virus replication obtained following transfection with wild-type viral DNA. By transcomplementation of a pol gene mutant in LMH cells, we were able to produce sufficient virus with the mutant genome to investigate the role of polymerase in covalently closed circular DNA amplification. Our results substantiate the hypothesis that covalently closed circular DNA is synthesized by the viral reverse transcriptase.
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PMID:Efficient duck hepatitis B virus production by an avian liver tumor cell line. 235 24

Nucleocapsid-pol fusion proteins have been detected by serological screening hepatocellular carcinoma tissues that contain hepatitis B virus (HBV) DNA. The existence of these fusion proteins suggests that HBV may synthesize its reverse transcriptase in a fashion analogous to the way that retroviruses synthesize and process a precursor. The accumulation of HBV reverse transcriptase intermediates in tumorous tissues and not in other tissues may be related to the absence of viral core particles and possibly contributes to tumor development.
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PMID:Putative reverse transcriptase intermediates of human hepatitis B virus in primary liver carcinomas. 241 1

Recent studies suggest that hepatitis B virus (HBV), despite being a DNA virus, replicates via an RNA intermediate (R. H. Miller, P. L. Marion, and S. W. Robinson, Virology 139:64-72, 1984; J. Summers and W. S. Mason, Cell 29:403-415, 1982). The HBV life cycle is therefore a permuted version of the RNA retroviral life cycle. Sequence homology between retroviral reverse transcriptase and the putative HBV polymerase gene product suggests the presence of an HBV reverse transcriptase (H. Toh, H. Hajashida, and T. Miyata, Nature (London) 305:827-829, 1983). As yet, there has been no direct evidence that reverse transcriptase activity is present in the viral particle. We used activity gel analysis to detect the in situ catalytic activities of DNA polymerases after sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Our studies demonstrated that HBV-like particles secreted by a differentiated human hepatoma cell line transfected with genomic HBV DNA contain two major polymerase activities which migrate as approximately 90- and approximately 70-kilodalton (kDa) proteins. This demonstrated, for the first time, that HBV-like particles contain a novel DNA polymerase-reverse transcriptase activity. Furthermore, we propose that the 70-kDa reverse transcriptase may be produced by proteolytic self-cleavage of the 90-kDa precursor protein.
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PMID:Two proteins with reverse transcriptase activities associated with hepatitis B virus-like particles. 244 93


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