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:2.7.7.48 (transcriptase)
9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Lipoprotein lipase (LPL), an enzyme which hydrolyzes triglycerides and participates in the catabolism of remnant lipoproteins, plays a crucial role in energy and lipid metabolism. The goal of this study was to analyze the expression and regulation of the LPL gene in human adrenals. Reverse transcriptase-polymerase chain reaction amplification and sequence analysis demonstrated the presence of LPL mRNA in fetal and adult human adrenal cortex. Furthermore, the human adrenocortical carcinoma cell line, NCI-H295, expresses LPL mRNA and protein, which is localized to the outer cellular membrane as demonstrated by immunofluorescence confocal microscopy and can be released in the medium by heparin addition. To asses whether the LPL gene is regulated by agents regulating adrenal steroidogenesis, NCI-H295 cells were treated with activators of second messenger systems. Whereas the calcium-ionophore A23187 did not affect LPL gene expression, treatment with phorbol 12-myristate 13-acetate decreased LPL mRNA levels in a time- and dose-dependent manner. This decrease after phorbol 12-myristate 13-acetate was associated with diminished heparin-releasable LPL mass and activity in the culture medium. Addition of the cAMP analog 8-Br-cAMP to NCI-H295 cells resulted in a rapid, but transient dose-dependent induction of LPL mRNA. Treatment with the protein synthesis inhibitor cycloheximide gradually induced, whereas simultaneous addition of cAMP and cycloheximide superinduced LPL mRNA levels. Nuclear run-on analysis indicated that the effects of cAMP and cycloheximide occurred at the transcriptional and post-transcriptional level, respectively. Transient co-transfection assays demonstrated that the first 230 base pairs of the proximal LPL promoter contain a cAMP-responsive element activated by protein kinase A and transcription factors belonging to the CREB/CREM family. These data indicate that LPL is expressed in human adrenal cortex and regulated in NCI-H295 adrenocortical carcinoma cells by activators of the protein kinase A and protein kinase C second messenger pathways in a manner comparable to P450scc, which catalyzes the first step in adrenal steroidogenesis. These observations suggest a role for LPL in adrenal energy and/or lipid metabolism and possibly in steroidogenesis.
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PMID:Expression and regulation of the lipoprotein lipase gene in human adrenal cortex. 866 37

The absorption of water and electrolytes by the gallbladder seems to be largely dependent upon a Na+/H+ exchange at the apical membrane of the gallbladder epithelium. To find out if the exchanger involved is the NHE3 isoform, as in other absorbing epithelia, two studies were performed using the rabbit gallbladder. First, we studied 22Na absorption in Ussing chambers with Krebs buffer as a control solution, and in the presence of amiloride (100, 200 or 1000 microM), ethyl-isopropyl-amiloride (EIPA, 1 or 5 microM), or the phorbol ester, phorbol 12-myristate 13-acetate (PMA, 1 microM). A net mucosal-to-serosal Na+ flux was observed with control buffer. No inhibition of this net flux was observed with 5 microM EIPA, and the IC50 for amiloride was found to be 200 microM. PMA induced a reduction of absorption by 30% that was prevented by incubation with calphostin C. Resistance to amiloride and EIPA, and inhibition by PMA are consistent with the involvement of the NHE3 isoform. The second study involved reverse-transcriptase polymerase chain reaction (RT-PCR) of total gallbladder RNA, with two primers designed to amplify a 645-base-pair fragment from NHE3 mRNA. A cDNA fragment of the expected size was actually obtained from gallbladder RNA, while RT-PCR of RNA from the liver, which does not contain NHE3, gave negative results. A sequence of 492 nucleotides of the amplified product was determined, which was almost superimposable onto the known sequence of the corresponding fragment of rabbit NHE3. It is concluded that, in rabbit gallbladder, neutral NaCl absorption is, at least in part, dependent on the NHE3 isoform of the Na+/H+ exchanger.
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PMID:Role of the NHE3 isoform of the Na+/H+ exchanger in sodium absorption by the rabbit gallbladder. 877 28

Primary lung adenocarcinomas originate from the progenitor cells of peripheral airway cells. Alveolar type II cells and Clara cells are the major progenitor cells of peripheral airway cells. Alveolar type II cells produce a lipid-protein complex called surfactant, which contains surfactant proteins SP-A, SP-B, SP-C and SP-D. Phosphatidylcholine (PC) and phosphatidylglycerol (PG) are believed to be essential for the surfactant function. Clara cells also express SP-A, SP-B and SP-D but not SP-C. In this study we examined the properties of the cancer cells isolated from the pleural effusion of a patient with primary lung adenocarcinoma by analyzing lipids, proteins and mRNAs. The cancer cells, designated as LC117 cells, were isolated from the pleural effusion of a patient with primary lung adenocarcinoma. The percent distributions of [14C]-acetate incorporated into PC and PG in the cancer cells were 55.7 and 1.1%, respectively. The disaturated species in total PC was 46.2%. Immunoblotting analysis using anti-SP-D monoclonal antibody revealed that the pleural effusion from a patient with lung adenocarcinoma contained SP-D. We determined the concentrations of SP-A and SP-D by enzyme-linked immunosorbent assay. The pleural effusions from this patient and the media incubated with cancer cells exhibited significant levels of SP-D as well as SP-A. Reverse transcriptase-polymerase chain reaction demonstrated that the tumor cells expressed mRNAs for SP-C as well as the other surfactant proteins. The results demonstrate that tumor cells from lung adenocarcinoma express all of surfactant-associated proteins, indicating that LC117 cells originate from alveolar type II cells. This study indicates that the combination of analyses of lipids, proteins and mRNAs in the cancer cells isolated from pleural effusion is useful to understand the property of lung adenocarcinoma.
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PMID:Lipid analysis and surfactant-associated protein expression in lung adenocarcinoma cells from pleural effusion. 893 61

The alpha subunits of the laminin-binding integrins alpha 3 beta 1, alpha 6 beta 1, and alpha 7 beta 1 have homologous sequences and are similar in structure. Two cytoplasmic variants, A and B, have been identified for each of these alpha subunits, although the alpha 3B splice variant has been detected only at the mRNA level. We prepared a panel of mouse monoclonal antibodies specific for the A and B variants of the alpha 3 subunit to study their tissue distribution. Four monoclonal antibodies react with alpha 3A, one of which recognizes only the nonphosphorylated form; of the three anti-alpha 3B antibodies, one cross-reacts with alpha 6B. Reverse transcriptase-PCR analysis of various human tissues revealed the presence of alpha 3B mRNA in brain, heart, and skeletal muscle. Moreover, the alpha 3B protein was detected by immunoblotting in brain and heart tissue but not in skeletal muscle. In contrast, alpha 3A mRNA and protein were present in all tissues studied. Thus, the expression of alpha 3B in adult tissues is more restricted than that of alpha 3A. Immunohistochemical studies showed that in brain tissue, both variants are exclusively expressed on small blood-vessel endothelium, whereas in heart tissue their distribution patterns differ markedly. Although alpha 3A is strongly expressed on vascular smooth muscle cells, alpha 3B is detected only on endothelial cells of veins. Expression of the two variant forms of alpha 3 in K562 cells revealed that the ligand-binding specificities of alpha 3A beta 1 and alpha 3B beta 1 are identical: both bind human laminin-2 and -4, laminin-5, and laminins isolated from bovine kidney, but not bovine laminin-2 and -4, mouse laminin-1, or human fibronectin. In addition, adhesion mediated by both integrins is induced to the same extent by phorbol 12-myristate 13-acetate. The alpha 3A, but not the alpha 3B subunit, is phosphorylated; and phosphorylation of alpha 3A increases after phorbol 12-myristate 13-acetate stimulation. Thus, we found no differences between the adhesion functions of the A and B variants of alpha 3.
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PMID:The A and B variants of the alpha 3 integrin subunit: tissue distribution and functional characterization. 911 16

The immortalized human chondrocyte cell line C-20/A4 has the ability to produce superoxide constitutively at low levels of 5.4 x 10(-2) nmol/min/10(6) cells (S.E.M. = +/-0.5, n = 30) and at raised levels upon stimulation with ionomycin and phorbol 12-myristate 13-acetate. Priming and anti-priming effects of interleukin (IL)-1 beta and IL-4, respectively, are also demonstrated. Reverse transcriptase polymerase chain reaction (RT-PCR) amplification using oligonucleotide primers to components of the NADPH oxidase enzyme complex showed mRNA expression of p22-phox, p40-phox and p47-phox. Western blot analysis using polyclonal antisera indicated the presence of the p47-phox p67-phox polypeptide components. These results show that the C-20/A4 cells contain an NADPH oxidase-like complex, similar to that found in other cell types, which produces superoxide anions.
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PMID:Detection of protein and mRNA of various components of the NADPH oxidase complex in an immortalized human chondrocyte line. 918 52

Porcine articular chondrocytes have the capacity to release superoxide in response to the addition of the calcium ionophore ionomycin in a concentration-dependent manner. This activity was not stimulated by the addition of fMetLeuPhe or the kinase activator phorbol myristate acetate (PMA). However, this release of superoxide was inhibited by iodonium diphenyl (IDP), suggesting the involvement of NADPH oxidase. Reverse transcriptase polymerase chain reaction (RT-PCR) using oligonucleotides designed against the known sequences for the human phagocyte NADPH oxidase showed the expression of p22-phox, p40-phox, and p47-phox mRNA, while Western blot analysis of chondrocyte extracts using polyclonal antisera raised against the human phagocyte NADPH oxidase suggested the presence of the p67-phox polypeptide. These results suggest that porcine articular chondrocytes can release reactive oxygen species using a NADPH oxidase-like complex.
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PMID:Detection of superoxide and NADPH oxidase in porcine articular chondrocytes. 929 50

In response to dibutyryl cyclic AMP (dbcAMP) and all-trans retinoic acid (ATRA), HL60 cells differentiate into granulocyte-like cells. Membrane-associated phospholipase D (PLD) activity in response to guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) or phorbol myristate acetate (PMA) was upregulated by these treatments. Reverse transcriptase-polymerase chain reaction (RT-PCR) analyses revealed that both hPLD1a and hPLD1b mRNAs were expressed in HL60 cells and that their expression levels increased during differentiation. hPLD2 mRNA levels rose dramatically during differentiation. These results suggest that the PLD genes undergo changes in transcriptional regulation during granulocytic differentiation of HL60 cells.
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PMID:Increased mRNA expression of phospholipase D (PLD) isozymes during granulocytic differentiation of HL60 cells. 951 45

While there is considerable evidence for phosphate (Pi) reabsorption in the distal tubule, Pi transport and its regulation have not been well characterized in this segment of the nephron. In the present study, we examined Na+-dependent Pi transport in immortalized mouse distal convoluted tubule (MDCT) cells. Pi uptake by MDCT cells is Na+-dependent and, under initial rate conditions, is inhibited by phosphonoformic acid (41 +/- 3% of control), a competitive inhibitor of Na+-Pi cotransport. The transport system has a high affinity for Pi (Km = 0.46 mM) and is stimulated by lowering the extracellular pH from 7.4 to 6.4 and inhibited by raising the pH from 7.4 to 8.4. Exposure to Pi-free medium for 21 h increased Na+-Pi cotransport from 2.1 to 5.5 nmol/mg of protein/5 minutes (p < 0.05) while parathyroid hormone, forskolin, and phorbol 12-myristate 13-acetate failed to alter Pi uptake in MDCT cells. Reverse transcriptase polymerase chain reaction of MDCT cell RNA provided evidence for the expression of the Npt1 but not the Npt2 Na+-Pi cotransporter gene. However, preincubation of MDCT cells with Npt1 antisense oligonucleotide led to only 20% inhibition of Na+-Pi cotransport, suggesting that other Na+-Pi cotransporters are operative in MDCT cells. Indeed, we showed, by ribonuclease protection assay, that MDCT cells express the ubiquitous cell surface receptors for gibbon ape leukemia virus (Glvr-1) and amphoteric murine retrovirus (Ram-1) that also function as Na+-Pi cotransporters. In summary, we demonstrate that the pH dependence and regulation of Na+-Pi cotransport in MDCT cells is distinct from that in the proximal tubule and suggest that different gene products mediate Na+-Pi cotransport in the proximal and distal segments of the nephron.
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PMID:Na+ -phosphate cotransport in mouse distal convoluted tubule cells: evidence for Glvr-1 and Ram-1 gene expression. 955 59

The transport of lactate and pyruvate across membranes of vestibular dark cells (VDC) may be important under aerobic, ischemic or hypoxic conditions. This study addresses the questions whether VDC from the gerbil contain an H+/monocarboxylate- cotransporter (MCT) and in which membrane, apical or basolateral, MCT is located. Uptake of monocarboxylates into VDC was monitored in functional studies by measuring the cytosolic pH (pHi) and by measuring the pH-sensitive equivalent short circuit current (Isc). Subtypes of the functionally identified MCT which are present in vestibular labyrinth tissues were identified as transcripts by cloning and sequencing of reverse-transcriptase polymerase chain reaction (RT-PCR) products. Monocarboxylates but not dicarboxylates induced a transient acidification of pHi which was inhibited by 5 mM alpha-cyano-4-hydroxycinnamate (CHC) but not by 1 microM DIDS or 500 microM pCMBS. The initial rate of acidification induced by monocarboxylates was dose-dependent in the range between 1 and 20 mM. K(m) values were for pyruvate 1.3, acetate 3.7, L-lactate 3.8 and D-lactate 7.3 mM. Both apical and basolateral application of monocarboxylates caused a transient increase of Isc which was sensitive to 5 mM CHC. RT-PCR revealed the presence of transcripts for the MCT subtypes MCT1 and MCT2. The identity of transcripts was confirmed by sequence analysis. These observations suggest that VDC contain an MCT in their apical and basolateral membrane and that the vestibular labyrinth contains transcripts for the subtypes MCT1 and MCT2.
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PMID:Vestibular dark cells contain an H+/monocarboxylate- cotransporter in their apical and basolateral membrane. 956 48

Progestogen suppresses the progression of endometrial cancer and has an important effect on the secretory change of human endometrium. We characterized the progestogen-induced alterations of gene expression in a human endometrial-cancer cell line using a mRNA differential-display reverse-transcriptase-polymerase-chain-reaction (DDRT-PCR) method. After 5-day incubation of Ishikawa endometrial-cancer cells, with or without 100 nM medroxyprogesterone acetate (M PA), total RNA was isolated from confluent cells. We identified 8 candidate genes by mRNA differential display by screening up to approximately 3,000 mRNA species. Among these, 2 genes named T21A and T21B showed a decrease in mRNA by MPA treatment when analyzed by Northern blot. Nucleotide sequence showed that clone T21A was part of human mitochondrial short-chain enoyl-CoA hydratase cDNA. The other clone, T21B, showed no homology with any known nucleotide sequences. Northern-blot analysis using T21A and T21B clones as probes showed a decrease in mRNA in human endometrium from the luteal stage, with high serum estradiol and progesterone levels, as compared with that from the early follicular stage, with low serum estradiol and progesterone levels, and that from the pre-ovulatory stage with high serum estradiol and low progesterone levels. These findings suggest that mRNA DDRT-PCR could be used to identify the candidate genes regulated by progestogen in human endometrial cancer and in normal human endometrium.
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PMID:Messenger RNA differential display reverse-transcriptase-polymerase-chain-reaction analysis of a progestogen-suppressive gene in a human endometrial-cancer cell line. 972 4


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