Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.3.1.28 (chloramphenicol acetyltransferase)
 5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We report here for the first time direct injection of genes into fish muscle in vivo. Plasmids used contain either SV40 early promoter, rabbit beta-cardiac myosin heavy chain promoter, human MxA promoter or an artificial promoter, fused to a chloramphenicol acetyltransferase (CAT) or beta-galactosidase reporter gene. CAT assays revealed that most gene constructs were highly expressed. Histochemical analysis showed that beta-galactosidase was strongly expressed at the site of injection within muscle fibres. This method provides an excellent system for testing expression of gene constructs, including those of mammalian origin, in fish muscle in vivo and has the potential for fish vaccination.
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PMID:Strong expression of foreign genes following direct injection into fish muscle. 191 96

Human MxA protein accumulates in the cytoplasm of interferon-treated cells and inhibits the multiplication of several RNA viruses, including Thogoto virus (THOV), a tick-borne orthomyxovirus that transcribes and replicates its genome in the cell nucleus. The antiviral mechanism of MxA was investigated by using two alternative minireplicon systems in which recombinant viral ribonucleoprotein complexes (vRNPs) of THOV were reconstituted from cloned cDNAs. A chloramphenicol acetyltransferase reporter minigenome RNA was expressed either by T7 RNA polymerase in the cytoplasm of transfected cells or, alternatively, by RNA polymerase I in the nucleus. The inhibitory effect of MxA was studied in both cellular compartments by coexpressing wild-type MxA or TMxA, an artificial nuclear form of MxA. Our results indicate that both MxA proteins recognize the assembled vRNP rather than the newly synthesized unassembled components. The present findings are consistent with previous data which indicated that cytoplasmic MxA prevents transport of vRNPs into the nucleus, whereas nuclear MxA directly inhibits the viral polymerase activity in the nucleus.
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PMID:MxA GTPase blocks reporter gene expression of reconstituted Thogoto virus ribonucleoprotein complexes. 1059 Jan 50

We describe here a specific and sensitive assay for biologically active bovine type-I interferon (IFN) in an Mx/CAT reporter gene assay. The assay is based on Madin-Darby Bovine Kidney cells transfected with a plasmid, containing a human MxA promoter driving a chloramphenicol acetyltransferase (CAT) cDNA. CAT expression was quantified in a commercially available enzyme linked immunosorbant assay. The response to recombinant bovine INF-alpha(1) was dose dependent between 0.25 and 125.0 iu/ml and was shown to be specific for type-I IFN as no significant effect was seen with a number of other cytokines, including IFN-gamma. This Mx/CAT reporter assay also has advantages in terms of simplicity and reliability over conventional cytopathic effect reduction assays used to quantify the IFN activity in bovine samples. The Mx/CAT reporter assay was used successfully to measure trophoblast derived type-1 IFN activity (IFN-tau) in uterine flushings collected from pregnant cows. IFN-tau is the pregnancy recognition signal produced in ruminants by pre-implantation embryos and was shown to increase markedly between the 12th (0.7+/-0.14 iu/ml) and 18th (44085.0+/-14414.2 iu/ml) day of pregnancy. In contrast, IFN-tau activity remained basal (0.5-0.7 iu/ml) in inseminated non-pregnant animals. Duplicate samples analysed using a cytopathic effect reduction assay correlated well (P<0.001; r(2)=0.945) with IFN levels obtained using the Mx/CAT reporter assay, confirming the reporter assay as a reliable substitute for the standard anti-viral IFN assay.
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PMID:Validation of an Mx/CAT reporter gene assay for the quantification of bovine type-I interferon. 1122 80

Alpha interferons (alpha-IFNs) are potent biologically active proteins synthesized and secreted by somatic cells during viral infection. Quantification of alpha-IFN concentrations in biological samples is used for diagnosis. More recently, recombinant IFNs have been used as antiviral, antiproliferative, and immunomodulatory therapeutic agents, and particularly for the treatment of chronic hepatitis C virus infection. For this purpose, IFN has recently been coupled to polyethylene glycol (PEG) to improve the pharmacokinetic properties. The measure of alpha-IFN in biological samples from treated patients could be useful to ensure compliance to therapy and the true IFN activity in relation to viral decay during follow-up. In particular, it could be used to monitor the PEG-IFN concentration in patients treated for hepatitis C virus infection. The most frequently used test is a bioassay based on the antiviral property of the IFN, but the assay is not highly reproducible. Here, we present a reporter test based on MxA promoter activation of chloramphenicol acetyltransferase expression (Mx-CAT). MxA is an antiviral protein induced and tightly regulated by alpha-IFN. The Mx-CAT assay showed good reproducibility of 15% and was suitable to quantify PEG-IFN and numerous other alpha-IFN subtypes as well, despite a differential MxA promoter activation in relation with the subtype. A good correlation was obtained with the reporter assay and a commercial enzyme-linked immunosorbent assay on samples from treated patients. This test could be useful for monitoring IFN therapy of chronically infected hepatitis C virus-infected patients treated with the standard IFN, PEG-IFN, and probably forthcoming recombinant IFNs.
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PMID:Quantification of different human alpha interferon subtypes and pegylated interferon activities by measuring MxA promoter activation. 1612 52