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

---

Query: EC:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

While the generalised pathway of collagen biosynthesis is well understood, the specific molecular interactions that drive chain recognition and assembly and the formation of tissue-specific extracellular supramolecular structures have not been elucidated. This review focuses on the use of in vitro collagen expression systems to explore some of these fundamental questions on the molecular basis of normal and mutant collagen assembly. Three in vitro expression/assembly systems are discussed. Firstly, a simple cell-free transcription/translation system to study the initial stages of collagen chain assembly. Secondly, a novel T7-driven high level expression system, using a recombinant vaccinia virus expressing T7 RNA polymerase, in transiently transfected cells which allows appropriate postranslational modification and collagen folding. Thirdly, the more complex questions of normal and mutant collagen extracellular matrix assembly are addressed by stable transfection and expression in cells which allow the formation of a 'tissue equivalent' matrix during long-term culture.
...
PMID:In vitro expression analysis of collagen biosynthesis and assembly. 950 70

Many studies using small-animal models suggest that angiotensin II (Ang II) plays an important role in neointimal formation after vascular injury. In the present study, we examined whether Ang II type 1 receptor (AT1)-mediated Ang II signaling is indispensable for the development of injury-induced neointimal formation using AT1a knockout (KO) mice. Reverse transcriptase-polymerase chain reaction analysis revealed that AT1 mRNA was not detectable in both uninjured and injured carotid arteries of KO mice, whereas the AT1 gene was expressed in uninjured carotid arteries of wild-type (WT) mice. At 14 days after injury, AT1 mRNA levels were increased by 1.5-fold in injured arteries of WT mice. Although AT2 mRNA was not detectable in uninjured arteries, expression of AT2 gene was induced in both animal groups at 2 weeks after injury. Vascular injury induced neointimal formation in KO mice as well as in WT mice. There were no significant differences between WT and KO mice in the extent of histological findings such as increased cross-sectional areas of the neointima and the media, the number of proliferating smooth muscle cells, and the amount of collagen and fibronectin. Treatment with subpressor doses of Ang II after injury enhanced the growth of neointima in WT mice but not in KO mice. Moreover, treatment with the selective AT1 antagonist CV-11974 before injury significantly decreased the formation of neointima in only WT mice, whereas treatment with the selective AT2 antagonist PD-123319 before injury had no effects in both animal groups. These results suggest that AT1-mediated Ang II signaling is not essential for the development of neointimal formation, although it may modify it.
...
PMID:Vascular injury causes neointimal formation in angiotensin II type 1a receptor knockout mice. 993 49

The formation of capillaries during development and tissue repair is likely to involve active reorganization of the actin cytoskeleton, although few studies have addressed this issue. Here, we have utilized an in vitro model of capillary morphogenesis whereby human umbilical vein endothelial cells are suspended within three-dimensional type I collagen gels. The cells undergo dramatic morphogenic changes to develop capillary lumens, tubes, and networks over 72 h of culture. Western blots using cell extracts of these gels over this time frame were performed using antibodies directed to various proteins associated with the actin cytoskeleton. Three proteins showed altered expression during the time course, and they were gelsolin, which increased fivefold; vasodilator-stimulated phosphoprotein (VASP), which increased twofold; and profilin, which increased threefold in expression between the 24- and the 72-h time points. Reverse transcriptase-polymerase chain reaction and Northern blot analysis revealed a similar increase in mRNA expression of the three proteins. After the onset of network formation, the differentiated endothelial cells (dECs) undergoing capillary morphogenesis were removed from collagen gels at 48 h of culture to compare their properties with untreated endothelial cells (uECs). These dECs showed two- to threefold increased spontaneous migration in Boyden chamber assays compared to uECs. The dECs also displayed a prominent spindle-shaped morphology and the novel presence of intranuclear gelsolin compared to uECs when both cell types were replated on type I collagen-coated microwells and glass coverslips. These data suggest that increased gelsolin, VASP, and profilin expression may play an important role in the regulation of capillary tube and network formation in three-dimensional extracellular matrix.
...
PMID:Coordinate induction of the actin cytoskeletal regulatory proteins gelsolin, vasodilator-stimulated phosphoprotein, and profilin during capillary morphogenesis in vitro. 1032 50

Nitric oxide plays a significant but incompletely understood role in fibroblast function and cutaneous wound collagen synthesis; however, the participation of inducible nitric oxide synthase (iNOS) in gastrointestinal anastomotic healing has not been studied. Male Sprague-Dawley rats underwent single-layer left colonic anastomosis. Animals were killed at 24-hour intervals postoperatively and the anastomosis was excised. Parallel uninjured colon tissue samples were also analyzed. Reverse transcriptase-polymerase chain reaction confirmed the absence of iNOS messenger RNA in control colon and expression of the gene in anastomotic tissue on all study days. Northern hybridization demonstrated maximal iNOS messenger RNA transcription on day 1 with decreased levels on days 3 and 5. iNOS enzyme activity, measured biochemically by the conversion of [(3) H-arginine to [(3) H]-citrulline ex vivo, was also maximal on day 1 (7.35 +/- 1.34 pmol/mg protein/min [+/- standard error of the mean], n = 10) and decreased on days 3 (4.37 +/- 2.32 pmol/mg protein/min; n = 6) and 5 (2.80 +/- 0.92 pmol/mg protein/min; n = 6). Immunohistochemical staining demonstrated that (1) iNOS expression is confined to a discrete cell population in the region of the anastomosis containing inflammatory cells; (2) those cells assume a highly conserved position on the luminal edge of the proliferating scar; and (3) the iNOS-expressing cells are present throughout the fibroplastic phase of healing. To functionally assess the role of iNOS in colonic healing, rats were treated with a continuous intravenous infusion of S-methylisothiourea (a selective inhibitor of iNOS) at a dosage of 200 mg/kg/day for 5 days after anastomosis. There was a significantly reduced anastomotic bursting pressure in rats treated with the inhibitor as compared to rats treated with intravenous normal saline solution (108.4 +/- 13.2 mm Hg vs. 148.4 +/- 10.3 mm Hg; P <0.05). These results suggest that iNOS gene expression is induced during colonic anastomotic healing, that it is present through all phases of healing but is maximal through the inflammatory phase, and that iNOS activity is required for optimal anastomotic healing.
...
PMID:Expression and function of inducible nitric oxide synthase during rat colon anastomotic healing. 1055 65

We have used the yeast two-hybrid system to clone the protein that interacts with the BFCOL1 (binding factor of a type-I collagen promoter) zinc-finger transcription factor that was cloned previously as the factor that binds to the two mouse proximal promoters of the type-I collagen genes. We utilized as bait the N-terminal domain of BFCOL1 that includes the zinc-finger DNA-binding domain. One cDNA contained a potential open reading frame for a polypeptide of 392 amino acids and was identical to PTRF (polymerase I and transcript-release factor), which is involved in transcription termination of the RNA polymerase I reaction. Northern-blot analysis revealed that the pattern of mRNA expression was similar to that of the type-I collagen gene. In addition, we detected the mRNA expression only in a fibroblast cell line and two bone cell lines, but not in other blood and neuronal cell lines. Recombinant protein was shown to enhance the binding of BFCOL1 to its binding site in the mouse proalpha2(I) collagen proximal promoter in vitro. The transient-transfection experiment showed that PTRF had a suppressive effect on the mouse proalpha2(I) collagen proximal promoter activity. We speculate that PTRF might play a role in the RNA polymerase II reaction as well as that of RNA polymerase I.
...
PMID:PTRF (polymerase I and transcript-release factor) is tissue-specific and interacts with the BFCOL1 (binding factor of a type-I collagen promoter) zinc-finger transcription factor which binds to the two mouse type-I collagen gene promoters. 1072 1

We have examined the catabolism of the proteoglycans aggrecan, decorin and biglycan in fresh tendon samples and in explant cultures of tissue from the tensional and compressed regions of young and mature bovine tendons. A panel of well-characterized antibodies that recognize glycosaminoglycan or protein (linear or neoepitope) sequences was used to detect proteoglycans and proteoglycan degradation products that were both retained within the tissue and released into the culture medium. In addition, a reverse-transcriptase-mediated PCR analysis was used to examine the mRNA expression patterns of tendon proteoglycans and aggrecanases. The results of this study indicate a major role for aggrecanase(s) in the catabolism of aggrecan in bovine tendon. The study also provides a characterization of glycosaminoglycan epitopes associated with the proteoglycans of tendon, illustrating age-related changes in the isomers of chondroitin sulphate disaccharides that remain attached to the core protein glycosaminoglycan linkage region after digestion with chondroitinase ABC. Evidence for a rapid turnover of the small proteoglycans decorin and biglycan was also observed, indicating additional molecular pathways that might compromise the integrity of the collagen matrix and potentially contribute to tendon dysfunction after injury and during disease.
...
PMID:Catabolism of aggrecan, decorin and biglycan in tendon. 1092 42

An autosomal recessive disorder, generalized atrophic benign epidermolysis bullosa, is a rare form of nonlethal type junctional epidermolysis bullosa. It is associated not only with skin fragility but also with other unique clinical features including widespread atrophic skin changes, alopecia, reduced axillary and pubic hair, dysplastic teeth, and dystrophic nails. The majority of generalized atrophic benign epidermolysis bullosa cases are caused by mutations in the COL17A1 gene coding for type XVII collagen (or the 180 kDa bullous pemphigoid antigen). Another candidate gene for mutations in some forms of generalized atrophic benign epidermolysis bullosa is LAMB3 encoding the beta3 chain of laminin 5. This report documents compound heterozygosity for novel mutations in LAMB3 of a Japanese patient showing typical clinical features of generalized atrophic benign epidermolysis bullosa. One is an A-to-G transversion at the splice acceptor site of intron 14, which is designated as a 1977-2A-->G mutation; the other is a deletion of 94 bp located at the junction of intron 18 and exon 19, which is a 2702-29del94 mutation. Reverse transcriptase polymerase chain reaction analysis suggested skipping of exon 19 in LAMB3 mRNA produced from the allele with 2702-29del94 and impaired stability of the aberrant mRNA transcribed from the second allele with the 1977-2A-->G mutation.
...
PMID:Compound heterozygosity for a point mutation and a deletion located at splice acceptor sites in the LAMB3 gene leads to generalized atrophic benign epidermolysis bullosa. 1095 Dec 52

The sequence of canine COL1A1 cDNA was determined from four overlapping COL1A1 RT-PCR products generated from canine fibroblast RNA. In the translated region, nucleotide identity between canine and human COL1A1 cDNA was 93.2%, although the canine sequence lacked nucleotides 204 to 215 in the region coding for the N-propeptide. Amino acid identity was 97.7%. Total RNA and type I collagen were collected from cultured skin fibroblasts of a 12-week-old male golden retriever with pathologic fractures suggestive of osteogenesis imperfecta (OI) and dentinogenesis imperfecta. Sequential, overlapping approximately 1,000-bp fragments of COL1A1 and COL1A2 cDNA were each amplified by RT-PCR using primers containing 5' T7 polymerase sites. These PCR products were transcribed with T7 RNA polymerase, hybridized into RNA duplexes, and cleaved at mismatch sites with RNase. The proband had an unique cleavage pattern for the fragment of COL1A1 mRNA spanning nucleotides 709 to 1,531. Sequence analysis identified a G to C point mutation for nucleotide 1,276, predicting a codon change from glycine (GGA) to alanine (GCA) for amino acid 208. This change disrupts the normal Gly-X-Y pattern of the collagen triple helix. Restriction enzyme digestion of the RT-PCR product was consistent with a heterozygous COL1A1 mutation. Type I collagen was labeled with 3H-proline, salt precipitated, and analyzed by SDS-PAGE. Pepsin digested alpha chains were over-hydroxylated, and procollagen processing was delayed. Thus, canine and human OI appear homologous in terms of clinical presentation, etiology, and pathogenesis.
...
PMID:Sequence of normal canine COL1A1 cDNA and identification of a heterozygous alpha1(I) collagen Gly208Ala mutation in a severe case of canine osteogenesis imperfecta. 1114 34

This study examines endothelin-induced modulation of extracellular matrix synthesis and remodeling by fibroblasts, and its potential role in the pathogenesis of systemic sclerosis (scleroderma). Endothelin-1 promoted fibroblast synthesis of collagen types I and III, but not fibronectin, by a mechanism dependent upon both ETA and ETB receptors. Conversely, endothelin-1 inhibited both protein expression of matrix metalloproteinase 1 and zymographic activity exclusively via ETA receptors. A dual regulatory role for endothelin-1 in transcriptional regulation was suggested by the ability of endothelin-1 to enhance steady-state levels of collagen mRNA and activate the proalpha2(I) collagen (Col1a2) promoter, but in contrast to reduce matrix metalloproteinase 1 transcript expression and suppress transcription of a human matrix metalloproteinase 1 promoter reporter construct in transient transfection assays. Although endothelin-1 significantly enhanced remodeling of three-dimensional collagen lattices populated by normal fibroblasts, this was not observed for lattices populated by systemic sclerosis fibroblasts. Promotion of matrix remodeling was dependent upon ETA receptor expression and was blocked by specific inhibitors of tyrosine kinases or protein kinase C. Reverse transcriptase polymerase chain reaction, S1 nuclease, and functional cell surface binding studies showed that normal and systemic sclerosis fibroblasts express both ETA and ETB receptors (predominantly ETA), but that ETA receptor mRNA levels and ETA binding sites on fibroblasts cultured from systemic sclerosis skin biopsies are reduced by almost 50%. Endothelin-1 is thus able to induce a fibrogenic phenotype in normal fibroblasts that is similar to that of lesional systemic sclerosis fibroblasts. Moreover, reduced responsiveness to exogenous endothelin-1 in systemic sclerosis suggests that downstream pathways may have already been activated in vivo. These data further implicate dysregulated endothelin-receptor pathways in fibroblasts in the pathogenesis of connective tissue fibrosis.
...
PMID:Fibroblast matrix gene expression and connective tissue remodeling: role of endothelin-1. 1123 16

Previously, we showed that exposure of human osteoblasts to titanium particles stimulates protein tyrosine phosphorylation (PTP), activates the transcription factor nuclear factor kappaB (NF-kappaB), and causes an approximately 50% decrease in the steady-state messenger RNA (mRNA) level of procollagen alpha1[I]. In this study, we identify three NF-kappaB binding sites within the human procollagen alpha1[I] gene promoter, show that titanium particles stimulate their binding of the NF-kappaB subunits Rel A (p65) and NF-kappaB1 (p50), and find NF-kappaB activation correlates with collagen gene suppression by titanium particles in osteoblasts. Protein tyrosine kinase (PTK) inhibitors, which significantly reduce the suppressive effect of titanium particles on collagen gene expression, inhibited NF-kappaB binding activity showing that titanium particle stimulation of PTK signals in osteoblasts are critical for both NF-kappaB activation and collagen gene expression. The antioxidant pyrrolidine dithiocarbamate (PDTC), which also inhibits the titanium particle suppression of collagen, abrogated the titanium particle activation of NF-kappaB, suggesting the involvement of redox signals in NF-kappaB-mediated collagen gene expression. The RNA polymerase II inhibitor actinomycin D (Act D) decreased procollagen alpha1[I] mRNA expression and effectively blocked the titanium-induced suppressive effect, suggesting that titanium particles activate a cascade of signals in osteoblasts, which result in a suppression of procollagen alpha1[I] mRNA. Collectively, these results show that titanium particles can activate NF-kappaB signaling in osteoblasts and suggest that NF-kappaB binding to the collagen gene promoter has a functional role in the down-regulation of procollagen alpha1[I] gene transcription.
...
PMID:Down-regulation of procollagen alpha1[I]] messenger RNA by titanium particles correlates with nuclear factor kappaB (NF-kappaB) activation and increased rel A and NF-kappaB1 binding to the collagen promoter. 1127 68


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---