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:3.2.1.31 (beta-glucuronidase)
7,680 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Zinc finger transcription factors (TFs(ZF)) were designed and applied to transgene and endogenous gene regulation in stably transformed plants. The target of the TFs(ZF) is the Arabidopsis gene APETALA3 (AP3), which encodes a transcription factor that determines floral organ identity. A zinc finger protein (ZFP) was designed to specifically bind to a region upstream of AP3. AP3 transcription was induced by transformation of leaf protoplasts with a transformation vector that expressed a TF(ZF) consisting of the ZFP fused to the tetrameric repeat of herpes simplex VP16's minimal activation domain. Histochemical staining of beta-glucuronidase (GUS) activity in transgenic AP3GUS reporter plants expressing GUS under control of the AP3 promoter was increased dramatically in petals when the AP3-specific TF(ZF) activator was cointroduced. TF(ZF)-amplified GUS expression signals were also evident in sepal tissues of these double-transgenic plants. Floral phenotype changes indicative of endogenous AP3 factor coactivation were also observed. The same AP3-specific ZFP(AP3) was also fused to a human transcriptional repression domain, the mSIN3 interaction domain, and introduced into either AP3GUS-expressing plants or wild-type Arabidopsis plants. Dramatic repression of endogenous AP3 expression in floral tissue resulted when a constitutive promoter was used to drive the expression of this TF(ZF). These plants were also sterile. When a floral tissue-specific promoter from APETALA1 (AP1) gene was used, floral phenotype changes were also observed, but in contrast the plants were fertile. Our results demonstrate that artificial transcriptional factors based on synthetic zinc finger proteins are capable of stable and specific regulation of endogenous genes through multiple generations in multicellular organisms.
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PMID:Heritable endogenous gene regulation in plants with designed polydactyl zinc finger transcription factors. 1227 Nov 25

Designer zinc finger transcription factors (TFs(ZF)) have been developed to control the expression of transgenes and endogenous genes in mammalian cells. Application of TFs(ZF) technology in plants would enable a wide range of both basic and applied studies. In this paper, we report the use of TFs(ZF) to target a defined 18-bp DNA sequence to control gene expression in plant cells and in transgenic plants. A beta-glucuronidase reporter gene was activated by using the designed six-zinc finger protein 2C7 expressed as a fusion with the herpes simplex virus VP16 transcription factor activation domain. Reporter gene expression was activated 5- to 30-fold by using TFs(ZF) in BY-2 protoplasts, whereas expression was increased as much as 450 times in transgenic tobacco plants. Use of a phloem-specific promoter to drive expression of the TFs(ZF) resulted in activation of the reporter gene in vascular tissues. Transgenic tobacco plants that produce 2C7 transcription factors were phenotypically normal through two generations, suggesting that the factors exerted no adverse effects. This study demonstrates the utility of zinc finger technology in plants, setting the stage for its application in basic and applied agricultural biotechnology.
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PMID:Regulation of transgene expression in plants with polydactyl zinc finger transcription factors. 1227 Nov 38

Studies of the human IFN-alpha subtype system have been hampered by the lack of efficient procedures to quantify and differentiate the expression of the highly homologous IFN-alpha subtypes. Here we evaluate four novel real-time PCR assays for the specific detection and quantification of IFN-alpha mRNA for the subtypes alpha(2), alpha(6), alpha(8) and alpha(1/13) in a combined assay in human peripheral blood mononuclear cells (PBMC). This included (a) the selection of beta-glucuronidase (GUS) as a suitable housekeeping gene for relative quantification; (b) verification of the specificity by using human DNA of different IFN-alpha subtypes; and (c) comparison of the amplification efficiencies among the different assays. This highly sensitive method allows the detection of low-level, constitutive IFN-alpha mRNA and shows differences in the composition of constitutive IFN-alpha subtypes compared to other cell types (HeLa and HEp-2). The in vitro stimulation of PBMC with Newcastle disease virus (NDV), Respiratory syncytial virus (RSV) or an inactivated Herpes simplex (HSV) preparation leads to the transcriptional induction of all IFN-alpha subtypes investigated but to different expression levels. Among the subtypes detected, IFN-alpha(13/1) and alpha(2) are the major transcripts followed by alpha(8), and finally alpha(6) as a minor transcribed subtype. Time-kinetics of IFN-alpha transcriptional activation also revealed variations in the course of IFN-alpha transcription between NDV, RSV or HSV. The data obtained from the real-time PCR assays correlated well with IFN-alpha(2) protein release. In conclusion, we have demonstrated the suitability and reliability of new real-time PCR assays for the rapid and efficient analysis of IFN-alpha subtype expression.
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PMID:Differential expression of IFN-alpha subtypes in human PBMC: evaluation of novel real-time PCR assays. 1273 74

A herpes simplex virus type 2 (HSV-2) UL24 beta-glucuronidase (UL24-betagluc) insertion mutant was derived from HSV-2 strain 186 via standard marker transfer techniques. Cell monolayers infected with UL24-betagluc yielded cytopathic effect with syncytium formation. UL24-betagluc replicated to wild-type viral titers in three different cell lines. UL24-betagluc was not virulent after intravaginal inoculation of BALB/c mice in that all inoculated animals survived doses up to 400 times the 50% lethal dose (LD50) of the parental virus. Furthermore, few UL24-betagluc-inoculated mice developed any vaginal lesions. Intravaginal inoculation of guinea pigs with UL24-betagluc at a dose equivalent to the LD50 of parental virus (approximately 5 x 10(3) PFU) was not lethal (10/10 animals survived). Although genital lesions developed in some UL24-betagluc-inoculated guinea pigs, both the overall number of lesions and the severity of disease were far less than that observed for animals infected with parental strain 186.
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PMID:Herpes simplex virus type 2 UL24 gene is a virulence determinant in murine and guinea pig disease models. 1605 42

Previous gene transfer studies of the herpes simplex virus type 1 (HSV-1) using the latency-associated transcript (LAT) promoter have reported a decrease in transgene expression in the brain over time, but the extent of this decrease has not been measured and it is unknown if expression eventually stabilizes. We examined LAT promoter-mediated transgene expression in the mouse brain for 1 year following intracranial injection with a HSV-1 vector expressing human beta-glucuronidase (GUSB). The vector genome copy number remained stable from 2 to 52 weeks. Quantitative reverse transcriptase PCR detected a peak of LAT intron expression at 2 weeks (corresponding to the end of the acute phase of viral infection), followed by stable expression during latency (13-52 weeks). The number of GUSB-positive cells also had a peak in the acute phase and then was stable during latency (13-52 weeks). GUSB enzymatic activity was maintained at 11% of normal at 6 and 12 months, indicating that the LAT promoter is capable of driving stable transgene expression in the brain.
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PMID:Stable levels of long-term transgene expression driven by the latency-associated transcript promoter in a herpes simplex virus type 1 vector. 1612 87

We have developed a plant virus-mediated transgene activation (VMTA) system that utilizes a viral expression vector to present the inducer. The concept was tested using two well characterized components: (i) an artificial promoter based on the yeast GAL4 upstream activating sequence and the minimal TATA element of Cauliflower Mosaic Virus 35S RNA promoter, and (ii) a transcriptional activator (TA) consisting of a fusion between the GAL4 DNA binding domain and the Herpes simplex virus VP16 activation domain. The TA was expressed under the control of the subgenomic promoter of a Tobacco Mosaic Virus-based expression vector. The VMTA system was functional in transient Agroinfiltration assays with the reporter gene beta-glucuronidase, the intracellular domain of the diabetes associated autoimmune antigen, IA-2ic, and with the anti-tetanus antibody 9F12. Transgenic lines harboring the reporter gene were also examined. The VMTA system displayed tight transcriptional control in both transient assays and in transgenic Nicotiana benthamiana plants carrying the TA-inducible reporter.
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PMID:Inducible expression in plants by virus-mediated transgene activation. 1620 7

We have inoculated a herpes simplex virus type 1 (HSV-1) vector into a variety of sites in the mouse brain and assayed the regions of latency and expression of a beta-glucuronidase (GUSB) cDNA from the latency-associated transcript promoter. Injection sites used were somatosensory cortex, visual cortex, striatum, dorsal hippocampus, and CSF spaces. Latent vector was detected in regions at a distance from the respective injection sites, consistent with axonal transport of vector. Regions of GUSB activity varied by injection site and included cerebral cortex, striatum, thalamus, hypothalamus, substantia nigra, hippocampus, midbrain, pons, medulla, cerebellum, and spinal cord. After a single injection, GUSB enzymatic activity reached wild-type levels in several brain regions. GUSB was found in some areas without any detectable vector, indicative of axonal transport of GUSB enzyme. GUSB-deficient mice, which have the lysosomal storage disease mucopolysaccharidosis (MPS) VII, have lysosomal storage lesions in cells throughout the brain. Adult MPS VII mice treated by injection of vector into a single site on each side of the brain had correction of storage lesions in a large volume of brain. The potential for long-term, widespread correction of lysosomal storage diseases with HSV-1 vectors is discussed.
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PMID:Widespread correction of lysosomal storage in the mucopolysaccharidosis type VII mouse brain with a herpes simplex virus type 1 vector expressing beta-glucuronidase. 1651 90

In mammals, the aryl hydrocarbon receptor (AhR) mediates expression of certain genes, including CYP1A1, in response to exposure to dioxins and related compounds. We have constructed a mouse AhR-mediated gene expression systems for a beta-glucuronidase (GUS) reporter gene consisting of an AhR, an AhR nuclear translocator (Arnt), and a xenobiotic response element (XRE)-driven promoter in transgenic tobacco plants. On treatment with the AhR ligands 3-methylcholanthrene (MC), beta-naphthoflavone (betaNF), and indigo, the transgenic tobacco plants exhibited enhanced GUS activity, presumably by inducible expression of the reporter gene. The recombinant AhR (AhRV), with the activation domain replaced by that of the Herpes simplex virus protein VP16, induced GUS activity much more than the wild-type AhR in the transgenic tobacco plants. Plants carrying AhRV expressed the GUS reporter gene in a dose- and time-dependent manner when treated with MC; GUS activity was detected at 5 nM MC on solid medium and at 12 h after soaking in 25 microM MC. Histochemical GUS staining showed that this system was active mainly in leaf and stem. These results suggest that the AhR-mediated reporter gene expression system has potential for the bioassay of dioxins in the environment and as a novel gene expression system in plants.
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PMID:Aryl hydrocarbon receptor (AhR)-mediated reporter gene expression systems in transgenic tobacco plants. 1787 99

To specify mechanisms of pathology development, we studied the activity of lysosomal glycosides in rabbit ocular tissues in experimental ophthalmoherpes. Rabbits with herpetic kepatitis show activation of beta-glucuronidase, beta-glycosidase, alpha-mannosidase in cornial epithelium and stroma, iris, aqueous humor of herpes-infected and contralateral eye. The activity of the above three glycosidases in the tears of children with herpetic keratitis was enhanced. If their activity does not regress after the treatment, ophthalmoherpes recurrence may be expected in the near future. For realization of its genetic program herpes simplex virus in infected cells activates synthesis of acid glycosidases this leading to degradation of cornial cell membranes, virus spread in the tissues and release of lysosomal enzymes into tear fluid.
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PMID:[Pathogenesis of ophthalmoherpes basing on the analysis of enzymes of the eye tissue and fluids]. 1872 Jul 21

Dioxin residues widely contaminate soil and agricultural products at low concentrations and may accumulate in organisms at the top of food chains owing to their physicochemical properties. In this study, we have developed novel, dioxin-inducible, reporter gene expression systems regulated by recombinant aryl hydrocarbon receptors (AhRs). The recombinant AhRs, referred to as XDVs, consist of the DNA-binding domain of the bacterial repressor protein LexA, a 90-kDa heat shock protein- and ligand-binding regulatory domain from mouse AhR, and the transactivation domain of herpes simplex virus regulatory protein VP16. Transgenic tobacco plants carrying XDVs absorb various AhR ligands, including 3-methylcholanthrene, beta-naphthoflavone and indigo from solid medium and vermiculite, and show dose- and time-dependent expression of the beta-glucuronidase reporter gene. The results clearly suggest that XDVs are functional transcription factors that respond to AhR ligands, and that the XDV-mediated reporter gene expression system is applicable to bioassays for dioxin residues in the environment.
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PMID:Recombinant aryl hydrocarbon receptors for bioassay of aryl hydrocarbon receptor ligands in transgenic tobacco plants. 1905 10


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