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:3.2.1.23 (beta-galactosidase)
14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Retinoic acid (RA) receptors (RARs) are ligand-inducible transcription factors that bind to specific DNA sequences associated with the regulatory regions of RA-regulatable genes. Since RA has been implicated as an important factor both in normal development and in teratological studies, one would like to have a model system that detects the presence of activated receptors during development. We have constructed a recombinant reporter gene that has three copies of the RA response element (RARE) from the RAR beta-2 promoter 5' to the herpes simplex virus thymidine kinase promoter; this regulatory region is coupled to the bacterial beta-galactosidase reporter gene. This construct was RA inducible in transient transfection assays in F9 embryonal carcinoma cells. Transgenic embryos with this reporter gene construct exhibited restricted and reproducible patterns of beta-galactosidase activity during embryogenesis, beginning between gestational ages day 7.5 and 8.5. At day 8.5, beta-galactosidase activity was detected in the closed neurotube and somites. Day 8.5 embryos, from pregnant females fed RA 14 hr earlier, exhibited a greater intensity and distribution of beta-galactosidase activity. Similarly, at later stages of gestation, maternal RA exposure resulted in enhanced embryonic beta-galactosidase expression. This type of transgenic indicator mouse should be useful in detailing the role of activated RARs during embryonic development.
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PMID:Transgenic indicator mice for studying activated retinoic acid receptors during development. 131 86

Recombinant DNA techniques were used to insert foreign genes into bovine herpesvirus-1 [infectious bovine rhinotracheitis virus (IBRV)] vectors which were attenuated by deletion and/or insertion mutations in the IBRV thymidine kinase (tk) gene. In one recombinant, the regulatory and coding sequences of the late pseudorabies virus (PRV) glycoprotein gIII gene, were inserted into the early IBRV tk gene. This recombinant efficiently expressed the PRV gIII gene indicating that immediate early IBRV proteins were competent to transactivate the late PRV gIII gene. IBRV vector viruses were also prepared in which the coding sequences of the early PRV tk gene, the late PRV gIII gene, and the E. coli beta-galactosidase gene were ligated to the late IBRV gIII promoter. Genotypes and phenotypes of the recombinant viruses were verified by restriction endonuclease and molecular hybridization experiments, thymidine plaque autoradiography, beta-gal plaque assays, and by immunoprecipitation experiments on extracts from 3H-mannose-labelled cells. The recombinant IBRV expressing beta-gal from the IBRV gIII promoter has been useful as an intermediate in the construction of IBRV vectors harboring foreign DNA sequences. The infectivity of the IBRV recombinant that expressed PRV gIII from the IBRV gIII promoter, was neutralized by polyclonal PRV antisera and by monoclonal antibodies to PRV gIII. The PRV gIII glycoprotein synthesized by the preceding recombinant has been used to coat microtiter test plate wells in a PRV gIII differential diagnostic test kit.
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PMID:Expression of porcine pseudorabies virus genes by a bovine herpesvirus-1 (infectious bovine rhinotracheitis virus) vector. 131 33

It is poorly understood why vaccines could not be developed for the control and prevention of African swine fever (ASF) virus infection. The aim of our study was to identify genes non-essential for ASF virus replication because there were indications that certain viral gene products, which apparently are non-essential for viral replication, conferred protection from death due to ASF. A cosmid library representing the genome of ASF virus strain France 64 was established and characterized. Then, in order to inactivate viral genes by insertion, the beta-galactosidase (beta-gal) gene was introduced either randomly or at specific locations of selected cloned DNA fragments. These constructions were transfected into cells which had been previously infected with a cell-culture-adapted viral strain in order to allow the generation of recombinant progeny virus. Viable recombinant progeny was identified by at least one of the following means: (1) expression of beta-gal; (2) detection of beta-gal specific DNA by plaque hybridization, and (3) absence of a functional product of the inactivated gene. Presently, we are characterizing a recombinant virus with an insertionally inactivated thymidine kinase gene.
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PMID:Approaches to the identification of non-essential genes of African swine fever virus. 148 51

Homologous recombination is shown to be specifically induced in Vero cells by infection with African swine fever (ASF) virus. The frequency of recombination induced by ASF virus infection between cotransfecting plasmids is comparable to that found after infection with the prototype poxvirus, vaccinia virus. The induction of recombination is accompanied by replication of the plasmid templates in the ASF virus-infected cells. An ASF virus insertion/expression plasmid vector containing the Escherichia coli reporter gene beta-galactosidase (beta-gal) fused to a viral promoter sequence was constructed. Recombination between homologous sequences present in both the plasmid vector and the virus genome led to the generation of recombinant viruses expressing the beta-gal gene. Visual screening of beta-gal+ plaques allowed the isolation and plaque purification of recombinant ASF viruses. The characterization of a beta-gal+ virus isolate showed that the beta-gal gene had been stably inserted into the thymidine kinase locus of the virus genome, thus demonstrating that controlled genetic manipulation of ASF virus can be achieved by homologous recombination in infected cells.
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PMID:Genetic manipulation of African swine fever virus: construction of recombinant viruses expressing the beta-galactosidase gene. 156 85

The stability and structure of the products of recombination in a fowlpox virus (FPV) system using the thymidine kinase (TK) gene as the insertion site were examined. A 4.6 kb chimeric DNA fragment from the pUV1 expression vector, containing the bacterial lacZ gene and the vaccinia virus P7.5 promoter, was ligated into the XbaI site of the FPV TK gene. The resulting vector, pFTKlacZb, was transfected into chicken embryo fibroblast cultures infected with FPV at an m.o.i. of 0.1. Recombinants were screened for the expression of beta-galactosidase. Five recombinants were isolated and plaque-purified to 80 to 90% for expression of beta-glucosidase. Serial cell culture passage of the recombinants led to the gradual reappearance of the non-recombinant parental phenotype. Southern hybridization analysis of EcoRI fragments from all five recombinants indicated that a single cross-over homologous recombination had occurred between either the 5' or the 3' end fragments of the TK gene, generating unstable intermediate recombinants incorporating the entire pFTKlacZb vector. Secondary intermolecular or intramolecular recombination of intergenic repetitive sequences within the intermediate recombinants appears to have resulted in frequent regeneration of the parental genotype and an infrequent generation of more stable recombinants. A method was developed to select stable recombinants by passage of the intermediate recombinants in chicken embryo fibroblast cultures treated with 5-bromo-2'-deoxyuridine.
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PMID:Structural analysis of unstable intermediate and stable forms of recombinant fowlpox virus. 165 7

The level of human thymidine kinase (TK) polypeptide is subject to cell cycle regulation. The enzyme is barely detectable in G1 phase but increases 10- to 20-fold by M phase. The low level of human TK in G1 phase is due primarily to the specific degradation of the protein during cell division. Substitution of heterologous promoters, removal of the introns, and deletion of all of the 3' untranslated region from the human TK gene do not affect cell cycle regulation of the enzyme. However, deletion of the carboxyl-terminal 40 amino acids or fusion of beta-galactosidase to the carboxyl terminus of human TK completely abolishes cell cycle regulation and stabilizes the protein throughout the cell cycle. These alterations do not significantly alter the specific enzymatic activity of TK. Changing the carboxyl terminus or deletion of the last 10 amino acids does not alter cell cycle regulation. These data demonstrate that residues near the carboxyl terminus of TK are essential for the cell cycle phase-specific degradation of the enzyme.
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PMID:Cell cycle regulation of thymidine kinase: residues near the carboxyl terminus are essential for the specific degradation of the enzyme at mitosis. 170 95

A genetically engineered herpes simplex virus type 1 (HSV-1, strain RH116) that expresses beta-galactosidase (beta-gal) was used as a marker to trace the route of interocular spread of HSV-1 after anterior chamber (AC) inoculation into BALB/c mice. Because RH116 is thymidine kinase deficient (TK-), the wild-type TK+ KOS strain of HSV-1 was used as a helper virus to complement RH116 during in vivo infection. After coinfection of BALB/c mice with RH116 and KOS in the AC of one eye, beta-gal expression by RH116 was detected in both the eyes and in the central nervous system (CNS). Our results suggest that after AC inoculation into BALB/c mice: (1) virus spreads from the injected eye to the CNS through parasympathetic fibers of the oculomotor nerve that supply the iris and ciliary body; (2) virus spread in the CNS is limited primarily to nuclei of the visual system and the suprachiasmatic area of the hypothalamus; and (3) virus is transmitted from the CNS to the retina of the contralateral eye by retrograde axonal transport through the optic nerve along the endocrine-optic pathway between the retina and the suprachiasmatic nucleus of the hypothalamus.
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PMID:Neural spread of herpes simplex virus after anterior chamber inoculation. 171 27

The ability to express recombinant genes in vivo offers potential new treatments for human disease if questions of safety and toxicity can be addressed. Complications of gene transfer could include, for example, overexpression of introduced genes for growth or angiogenic factors or insertional mutagenesis, both of which could cause uncontrolled cell growth. We report the development of a suicide retroviral vector that provides a method to eliminate cells undergoing rapid growth in vivo. A murine amphotropic retroviral vector was constructed in which the gene for herpesvirus thymidine kinase was included to render proliferating cells sensitive to ganciclovir, and the Escherichia coli beta-galactosidase gene served as a reporter. This vector's efficacy was first assessed in vitro, and beta-galactosidase activity was abolished in several cell lines after treatment with ganciclovir. In vivo, a transplantable murine CT26 adenocarcinoma whose cells were transduced with this vector regressed completely after administration of ganciclovir. In contrast, expression in nondividing cells within rabbit arteries transduced by retroviral infection in vivo was unaffected. This suicide vector therefore eliminates transformed cells but allows survival of normal nondividing cells that express its specific recombinant genes in vivo, and may thus improve the safety and efficacy of gene transfer into living organisms.
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PMID:Selective elimination of recombinant genes in vivo with a suicide retroviral vector. 175 52

The enzymic composition of 7 human mesothelioma lines propagated in nude mice was compared with 4 of the original and 15 additional mesotheliomas sampled during the patients' surgery. The xenografts exhibited several-fold higher thymidine kinase (TK), uridine kinase (UK), phosphoserine phosphatase (PSP) and peptidyl proline hydroxylase (PPH) concentrations than the fresh human samples, while their DNA, gamma-glutamyl transpeptidase (GGT) and beta-galactosidase (Bgal) contents remained similar. The volume growth rate of the xenografts (doubling time, DT = 9.23 +/- 1.25 days) was much faster than that of tumors in the human host, and the decline of this rate with increasing nodule size was accompanied by decreases in TK and PSP concentrations. This first quantitative biochemical study of xenografted human neoplasms indicates that 1) pleural mesotheliomas, though preserving their histological characteristics after heterotransplantation, show considerable increases of enzymes in nucleic acid, collagen, and nonessential amino acid synthesis, and that 2) the concentration of TK is a good indicator of the different growth properties of tumors in a mouse rather than in the human host.
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PMID:Enzymic composition and growth rate of human pleural mesothelioma transplants in nude mice. 176 9

In-frame codon insertion and deletion mutants were constructed in a plasmid containing the sequence that encodes ICP0, a transcriptional activator of herpes simplex virus type 1 (HSV-1). The effect of these mutations was analyzed in a transient expression assay using the promoters for, the IE-0 gene (an immediate early (alpha) gene), the thymidine kinase gene (an early (beta) gene), and the glycoprotein C gene (a late (gamma) gene) fused to reporter cassettes that encoded either beta-galactosidase or chloramphenicol acetyl transferase. Assays were performed in the presence or absence of a plasmid encoding ICP4, the major regulatory protein of HSV-1. Our results demonstrate that ICP0-mediated transactivation varied depending on the position of the insertion in the gene. One region of this protein was consistently shown to be required for full activation of each promoter examined either in the presence or in the absence of ICP4. This region overlaps with a cysteine-rich region and coincides with a transactivator domain identified in another extensive mutational analysis of this sequence. Analysis of the deletion mutants generated in this study demonstrated that the carboxy-terminal regions were required for activation in certain circumstances and that this varied depending on the promoter being assayed and the cell type in which the analysis was performed.
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PMID:Mutational analysis of the sequence encoding ICP0 from herpes simplex virus type 1. 184 23


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