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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mast cell activation requires Cl(-) flux, which maintains the driving force for entry of extracellular calcium and initiates release of mediators such as histamine. However, chloride channel expression in mast cells has been poorly understood. For the first time, reverse transcriptase-polymerase chain reaction shows that rat-cultured mast cells (RCMC) and peritoneal mast cells (PMC) contain mRNA for the cystic fibrosis transmembrane conductance regulator (CFTR), an important chloride channel. Immunostaining with an anti-CFTR antibody indicates expression of CFTR in PMC and RCMC. Mast cell CFTR is a functional Cl(-) channel because it is capable of mediating Cl(-) flux in response to elevated cAMP. An inhibitor of CFTR-dependent Cl(-) flux, diphenylamine-2-carboxylate down-regulates mast cell mediator release. These results show that rat mast cells express a functional CFTR, which might be important in mediator release.
...
PMID:Expression and functional characterization of CFTR in mast cells. 1178 80

Guanylyl cyclase C (GC-C) is a membrane-associated form of guanylyl cyclase and serves as the receptor for the heat-stable enterotoxin (ST) peptide and endogenous ligands guanylin, uroguanylin, and lymphoguanylin. The major site of expression of GC-C is the intestinal epithelial cell, although GC-C is also expressed in extraintestinal tissue such as the kidney, airway epithelium, perinatal liver, stomach, brain, and adrenal glands. Binding of ligands to GC-C leads to accumulation of intracellular cGMP, the activation of protein kinases G and A, and phosphorylation of the cystic fibrosis transmembrane conductance regulator (CFTR), a chloride channel that regulates salt and water secretion. We examined the expression of GC-C and its ligands in various tissues of the reproductive tract of the rat. Using reverse transcriptase and the polymerase chain reaction, we demonstrated the presence of GC-C, uroguanylin, and guanylin mRNA in both male and female reproductive organs. Western blot analysis using a monoclonal antibody to GC-C revealed the presence of differentially glycosylated forms of GC-C in the caput and cauda epididymis. Exogenous addition of uroguanylin to minced epididymal tissue resulted in cGMP accumulation, suggesting an autocrine or endocrine activation of GC-C in this tissue. Immunohistochemical analyses demonstrated expression of GC-C in the tubular epithelial cells of both the caput epididymis and cauda epididymis. Our results suggest that the GC-C signaling pathway could converge on CFTR in the epididymis and perhaps control fluid and ion balance for optimal sperm maturation and storage in this tissue.
...
PMID:Expression of the receptor guanylyl cyclase C and its ligands in reproductive tissues of the rat: a potential role for a novel signaling pathway in the epididymis. 1244 76

Following adeno-associated virus (AAV)-mediated transduction, cellular RNA preparations can be contaminated with AAV single-stranded DNA. The single-stranded DNA genome of recombinant AAV vectors can serve as an efficient, but undesirable, template for traditional reverse transcriptase-polymerase chain reaction (RT-PCR) methods. Consequently, recombinant AAV gene therapy presents a unique challenge to the design of sensitive and reliable methods to detect vector-derived mRNA. Several methods have been proposed to reduce the presence of single- and double-stranded vector DNA without compromising RNA specificity. For example, DNase I, although widely used, can be ineffective at completely removing the AAV single-stranded DNA genome. We have developed a sensitive real-time RNA-Specific reverse transcriptase PCR (RS-PCR) method that is independent of DNase I treatment. The RS-PCR method relies on the generation of a first-strand cDNA template using a primer with a linker sequence, X, at the 5'- end such that synthesis of second-strand cDNA incorporates the X-linker sequence into the cDNA template. The RS-PCR then utilizes forward and reverse primers targeting AAV vector sequence and the X-primer site, respectively, while a vector-specific Taqman probe makes sensitive real-time detection possible. We present data to validate the sensitivity and RNA specificity of the RS-PCR method and propose two unique endogenous control strategies by monitoring expression of both beta-glucuronidase and endogenous cystic fibrosis transmembrane conductance regulator (CFTR). Finally, we demonstrate the utility of this new RS-PCR method in detecting recombinant AAV-CFTR expression, including, an in vitro transduction assay and methods to support both preclinical and clinical trials.
...
PMID:A sensitive, real-time, RNA-specific PCR method for the detection of recombinant AAV-CFTR vector expression. 1293 41

The cystic fibrosis transmembrane conductance regulator (CFTR) is a cAMP-dependent protein kinase A-activated chloride channel that resides on the apical surface of epithelial cells. One unusual feature of this protein is that during biogenesis, approximately 75% of wild type CFTR is degraded by the endoplasmic reticulum (ER)-associated degradative (ERAD) pathway. Examining the biogenesis and structural instability of the molecule has been technically challenging due to the limited amount of CFTR expressed in epithelia. Consequently, investigators have employed heterologous overexpression systems. Based on recent results that epithelial specific factors regulate both CFTR biogenesis and function, we hypothesized that CFTR biogenesis in endogenous CFTR expressing epithelial cells may be more efficient. To test this, we compared CFTR biogenesis in two epithelial cell lines endogenously expressing CFTR (Calu-3 and T84) with two heterologous expression systems (COS-7 and HeLa). Consistent with previous reports, 20 and 35% of the newly synthesized CFTR were converted to maturely glycosylated CFTR in COS-7 and HeLa cells, respectively. In contrast, CFTR maturation was virtually 100% efficient in Calu-3 and T84 cells. Furthermore, inhibition of the proteasome had no effect on CFTR biogenesis in Calu-3 cells, whereas it stabilized the immature form of CFTR in HeLa cells. Quantitative reverse transcriptase-PCR indicated that CFTR message levels are approximately 4-fold lower in Calu-3 than HeLa cells, yet steady-state protein levels are comparable. Our results question the structural instability model of wild type CFTR and indicate that epithelial cells endogenously expressing CFTR efficiently process this protein to post-Golgi compartments.
...
PMID:Efficient intracellular processing of the endogenous cystic fibrosis transmembrane conductance regulator in epithelial cell lines. 1506 92

Interaction between the cystic fibrosis transmembrane conductance regulator (CFTR), a CAMP-activated Cl- channel, and epithelial Na+ channel (ENaC) has been proposed as the major mechanism regulating uterine fluid absorption and secretion. Differential expression of these ion channels may give rise to dynamic changes in the fluid environment affecting various reproductive events in the female reproductive tract. This study investigated the expression and localization of CFTR and ENaC during the pre-implantation period. Semi-quantitative reverse transcriptase polymerase chain reaction and immunohistochemistry were used to study the expression and localization of CFTR and ENaC in uteri collected from mature superovulated female mice. RT-PCR showed maximal ENaC and CFTR expression on day 3 after mating. Maximal immunoreactivity was also observed for both ENaC and CFTR on day 3 after mating. However, ENaC was immunolocalized to the apical membrane of both luminal and glandular epithelia, while CFTR was predominantly found in the stromal cells rather than the epithelial cells. Differential expression and localization of CFTR and ENaC provide a molecular mechanism by which maximal fluid absorption can be achieved immediately prior to implantation, to ensure the immobilization of the blastocyst necessary for implantation.
...
PMID:Differential expression and localization of CFTR and ENaC in mouse endometrium during pre-implantation. 1522 19

Airway submucosal gland serous cells express the cystic fibrosis transmembrane conductance regulator (CFTR) and secrete antimicrobial, anti-inflammatory, and antioxidant molecules. In cystic fibrosis, diminished gland secretion may impair innate airway host defenses. We used Calu-3 cells as a serous cell model to study the types of proteins released, the pathways that release them, and the possible involvement of CFTR activity in protein release. Many proteins were secreted constitutively into the apical fluid and showed increased release to agonists. We identified some of them by high pressure liquid chromatography-mass spectrometry and reverse transcriptase PCR, including lysozyme, siderocalin (the protein NGAL), which inhibits bacterial growth by binding iron-containing siderophores, HSC-71, which is thought to have anti-inflammatory properties, and the serine protease inhibitors alpha-1-antitrypsin and alpha-1-antichymotrypsin, which may function as antimicrobials as well as play a potential role in diminishing the activation of epithelial Na(+) channels by serine proteases. We used an enzyme-linked immunosorbent assay to quantify lysozyme secretion by Calu-3 cells in response to various agonists and inhibitors. Forskolin increased the lysozyme secretion rate (J(lyz)) from 32 to 77 ng/hr/cm(2) (n = 36, p < 0.005). Thapsigargin increased J(lyz) from 40 to 63 ng/h/cm(2) (n = 16, p < 0.005), and forskolin plus thapsigargin further increased the forskolin-stimulated J(lyz) by 48% (n = 9, p < 0.05). 1-Ethyl-benzimidazolinone and carbachol were less effective. Glibenclamide inhibited basal and stimulated J(lyz), but clotrimazole was without effect. CFTR(inh)172 caused a small (15%) but significant inhibition of forskolin-stimulated J(lyz) without affecting basal J(lyz). Thus, Calu-3 cells secrete diverse proteins that in aggregate would be expected to suppress microbial growth, protect the airways from damage, and limit the activation of epithelial Na(+) channels via serine proteases.
...
PMID:Regulation of antiprotease and antimicrobial protein secretion by airway submucosal gland serous cells. 1523 67

The cystic fibrosis transmembrane conductance regulator (CFTR) is one of the most intensively investigated Cl- channels. Different mutations in the CFTR gene cause the disease cystic fibrosis (CF). CFTR is expressed in the apical membrane of various epithelial cells including the intestine. The major organ affected in CF patients is the lung, but it also causes an important dysfunction of intestinal ion transport. The modulation of CFTR mRNA expression by atrial natriuretic peptide (ANP) was investigated in rat proximal colon and in human intestinal CaCo-2 cells by RNase protection assay and semi-quantitative reverse transcriptase PCR techniques. Groups of rats subjected to volume expansion or intravenous infusion of synthetic ANP showed respective increases of 60 and 50% of CFTR mRNA expression in proximal colon. CFTR mRNA was also increased in cells treated with ANP, reaching a maximum effect at 10(-9) M ANP, probably via cGMP. ANP at 10(-9) M was also able to stimulate both the CFTR promoter region (by luciferase assay) and protein expression in CaCo-2 cells (by Western blot and immunoprecipitation/phosphorylation). These results suggested the involvement of ANP, a hormone involved with extracellular volume, in the expression of CFTR in rat proximal colon and CaCo-2 intestinal cells.
...
PMID:Atrial natriuretic peptide modulates cystic fibrosis transmembrane conductance regulator chloride channel expression in rat proximal colon and human intestinal epithelial cells. 1661 90

Impaired glucose tolerance and overt diabetes mellitus are becoming increasingly common complications of cystic fibrosis (CF), most probably merely as a result of increased life expectancy. In order to understand the pathophysiology of cystic fibrosis-related diabetes (CFRD), knowledge on the possible expression and cell distribution of the cystic fibrosis transmembrane conductance regulator (CFTR) protein within the endocrine pancreas is required. In this report, we establish the first evidence for expression of CFTR protein in rat pancreatic islets by using independent techniques. First reverse transcriptase-polymerase chain reaction (RT-PCR) amplification showed that CFTR mRNA is present in isolated islets of Langerhans. Furthermore, the analysis of flow cytometry-separated islet cells indicated that the level of CFTR transcripts is significantly higher in the non-beta than in beta-cell populations. The expression of CFTR protein in rat islet cells was also demonstrated by Western blotting and the level of expression was also found significantly higher in the non-beta than in beta-cell populations. Last, in situ immunocytochemistry studies with two monoclonal antibodies recognizing different CFTR epitopes indicated that CFTR expression occurs mainly in glucagon-secreting alpha-cells.
...
PMID:Expression and localization of cystic fibrosis transmembrane conductance regulator in the rat endocrine pancreas. 1804 Aug 94

To verify the hypothesis that cystic fibrosis transmembrane conductance regulator (CFTR) is expressed in neurons of the human spinal cord, we investigated the presence and distribution of CFTR protein and mRNA in different segments of the human spinal cord obtained from autopsies. The techniques employed included reverse transcriptase-polymerase chain reaction (RT-PCR) to detect CFTR gene expression, in situ hybridization to detect mRNA distribution, and immunohistochemistry to detect protein distribution. The specificity of these experiments was established with extensive controls. We found widespread and abundant expression of CFTR in neurons of the human spinal cord. CFTR protein and mRNA are localized to the cytoplasm of neurons in all segments of the spinal cord but not to glial fibrillary acidic protein (GFAP)-positive cells. CFTR is a very important molecule, acting as a chloride channel and regulating many physiological functions, including salt transport, fluid flow, and intracellular ion concentrations. Its mutation causes cystic fibrosis. Our finding of abundant CFTR in the spinal cord suggests that this molecule may be significant in the normal function and pathology of the spinal cord.
...
PMID:Expression and distribution of cystic fibrosis transmembrane conductance regulator in neurons of the spinal cord. 1953 35

The importance of the molecule cystic fibrosis transmembrane conductance regulator (CFTR) is reflected in the many physiological functions it regulates. It is known to be present in epithelial cells of the lungs, pancreas, sweat glands, gut, and other tissues, and gene mutations of CFTR cause cystic fibrosis (CF). We studied the expression and distribution of CFTR in the human brain with reverse transcriptase polymerase chain reaction, in situ hybridization, and immunohistochemistry. This study demonstrates widespread and abundant expression of CFTR in neurons of the human brain. Techniques of double labeling and evaluation of consecutive tissue sections localized CFTR protein and mRNA signals to the cytoplasm of neurons in all regions of the brain studied, but not to glial cells. The presence of CFTR in central neurons not only provides a possible explanation for the neural symptoms observed in CF patients, but also may lead to a better understanding of the functions of CFTR in the human brain.
...
PMID:Expression and distribution of cystic fibrosis transmembrane conductance regulator in neurons of the human brain. 1965 4


<< Previous 1 2 3 Next >>

---