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)

12(R)-Hydroxy-5,8,14(Z,Z,Z)-eicosatrienoic acid (12(R)-HETrE) is an arachidonic acid metabolite formed by the corneal epithelium of several species, porcine leukocytes, and human and rat epidermal cells. It is a potent, stereospecific proinflammatory and angiogenic factor and its synthesis is increased manyfold in inflamed tissues, e.g. cornea and skin. It is possible that the angiogenic activity of 12(R)-HETrE is due to a direct mitogenic effect on microvessel endothelial cells via yet to be elucidated cellular and molecular mechanisms. In the present study, we demonstrated the ability of 12(R)-HETrE to stimulate the growth of quiescent endothelial cells in a time- and concentration-dependent manner with a maximal effect at 0.1 nM. This effect was highly stereospecific since its enantiomer, 12(S)-HETrE, had no effect within the same concentration range. Northern blot analysis and transient transfection experiments with chloramphenicol acetyltransferase constructs of oncogene promoter regions demonstrated significant increases over control (0.5% fetal calf serum) in c-myc-, c-jun, and c-fos mRNA levels and expression in cells treated with 0.1 nM 12(R)-HETrE. Electrophoretic mobility shift assay of nuclear protein extracts from cells treated with 12(R)-HETrE with specific radiolabeled oligonucleotides corresponding to known transcriptional binding sites, including AP-1, AP-2, SP1, TRE, NF kappa B, TFIID, OKT1, CREB, CTF/NF1, and GRE demonstrated a markedly rapid and specific increase in the binding activity of NF kappa B and to a lesser extent, AP-1. No significant increase was observed in the binding of other transcription factors assayed as compared to control (untreated) cells. Since the protooncogenes (c-fos, c-jun, and c-myc) are immediate early response genes that are implicated in the process of cell proliferation and differentiation, and activation of certain transcription factors, in particular NF kappa B, is associated with the immediate response of the cell to an injury, we propose that 12(R)HETrE's mitogenic and angiogenic activities are mediated, in part, via the activation of NF kappa B and expression of these protooncogenes.
 ...
PMID:Activation of nuclear factor kappa B and oncogene expression by 12(R)-hydroxyeicosatrienoic acid, an angiogenic factor in microvessel endothelial cells. 752 72

We have previously shown in a transgenic mouse line, in which 5.2 kb of the elastin promoter was linked to the reporter enzyme chloramphenicol acetyltransferase (CAT), that the highest levels of expression were found in embryonic lungs and aorta, while lower levels were detected in other elastin-containing tissues. Furthermore, in general, expression of the transgene showed developmental regulation similar to that of the endogenous gene. However, the precise location of cellular expression could not be determined in this model. To overcome this limitation, we have developed a similar model, but replaced CAT with the reporter enzyme beta-galactosidase. Enzyme activity was readily detected in the transgenic mouse embryos in expected regions of tissue forming elastic fibers, including the dermis and elastic cartilage. Of considerable interest, however, was the novel finding of expression in specific areas of neuroepithelium of the brain and in the perichondrium surrounding areas destined to form hyaline cartilage in endochondral bone formation. These latter areas included all the bones of the limbs, the spine and rib cage. It appeared that these segments of elastin expression demarcated the border between the developing cartilage and the surrounding mesenchymal tissue. Elastin promoter expression was also found in developing somites, in the mesenchymal layer of the forming cornea of the eye, in the genital tubercle and in the epithelium destined to form the olfactory epithelium. These findings indicate that the elastin promoter is activated during embryonic development in a variety of tissues, suggesting that elastin gene expression may play a role in organizing cutaneous, skeletal and neural structures.
 ...
PMID:Expression of the elastin promoter in novel tissue sites in transgenic mouse embryos. 1076 40

Interphotoreceptor retinoid binding protein (IRBP), a putative component of the visual cycle, is expressed selectively in the retina and pineal gland. This study examined whether site-specific DNA hypomethylation plays a role in this expression regulation. Southern blotting of HpaII and MspI digests of DNA from various bovine and murine tissues (whole brain, retina, pineal gland, superior colliculus, cortex, thymus, habenular nucleus, cornea, liver, tail, and kidney) revealed that specific CpG dinucleotides in the IRBP gene promoter are hypomethylated in DNA from retinal photoreceptor cells and pineal gland compared to DNA from other tissues. These sites are methylated in DNA from non-photoreceptor retinal cells. Exogenous methylation of these sites diminished DNA:protein binding in electrophoretic mobility shift assays. HpaII methylation of chloramphenicol acetyltransferase reporter constructs suppressed IRBP but not SV40 promoter activity in transiently transfected primary cultures of embryonic chick retinal cells. These data indicate that specific cytosines in the bovine and murine IRBP promoters are unmethylated in photoreceptive cells but methylated in other tissues. This differential DNA methylation may modulate IRBP gene expression since exogenous methylation of the murine sites suppresses reporter gene transcription, apparently by inhibiting DNA:protein binding events.
 ...
PMID:Site-specific DNA hypomethylation permits expression of the IRBP gene. 1113 9

Development of gene transfer methods that can precisely deliver therapeutic genes to the localized or targeted tissue(s) would be highly beneficial in developing new gene therapy approaches and may also extend animal models for studying in vivo gene function and regulation at molecular levels in the selected tissues. We investigated lipid- and AAV-mediated gene transfer in rabbit cornea using a lamellar flap-technique. The goals of this study were to (1) analyze methods for in situ gene transfer into keratocytes, (2) identify efficient and suitable vectors for gene transfer into keratocytes, and (3) characterize times of first detectable expression, localization and duration of transgene expression in keratocytes with different vectors. A lamellar flap was produced in the rabbit cornea with a microkeratome. Recombinant adeno-associated viral vector (rAAV) expressing either beta-galactosidase (rAAV-beta-gal) or chloramphenicol acetyltransferase (rAAV-CAT) reporter genes, or plasmid-cationic lipid complexes expressing CAT (pMP6-CAT) or beta-galactosidase (pTR-beta-gal) were applied beneath the lamellar flap for two minutes. The flap was repositioned and eyelids sutured overnight. Corneas were removed at 4hr, 12hr, 36hr, 3 days, 7 days, or 10 days after application and either fixed in 2% formaldehyde, cryosectioned and stained for beta-galactosidase activity or homogenized and measured for CAT levels by ELISA. Corneas infected with rAAV-beta-gal vector showed positive beta-gal staining in the center and periphery of the flap interface in whole corneas and corneal beds at 3, 7, and 10 days, but not at earlier time points. Corneas treated with pTR-beta-gal plasmid vector showed positive beta-gal expression at the interface at 4, 12 and 36hr, but not at 3 or 7 days. The posterior surface of the lamellar interface where the vector was applied showed more expression than the overlying anterior surface with both plasmid and viral vectors. The level of gene expression was less with plasmid vector than viral vector monitored using beta-gal staining. CAT-ELISA confirmed expression of the CAT reporter gene with either the plasmid or rAAV vector. These results demonstrate that foreign genes can be introduced into keratocytes with plasmid or viral vectors using a lamellar flap to gain access to the stroma. The expression profile of the reporter genes depended on the vector. Transfection of keratocytes with plasmid vectors produced rapid expression of the reporter genes, but for a short duration. Reporter gene expression following transduction by rAAV vector was delayed several days, but was at higher levels and for a longer duration. This is the first report to demonstrate selective gene transfer into keratocytes and would be highly useful in studying function and regulation of genes in vivo and may eventually furnish a tool for the treatment of corneal dystrophies.
 ...
PMID:Gene transfer into rabbit keratocytes using AAV and lipid-mediated plasmid DNA vectors with a lamellar flap for stromal access. 1257 66