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

Blastocyst formation and expansion are dependent on the differentiation and function of a proper transport of nutrients through the trophectoderm (TE) enclosing the inner cell mass (ICM). Coincident with compaction and cavitation, glucose becomes the preferred energy substrate of the early embryo. These hallmarks in early development require well-orchestrated gene expression patterns specifically with regard to timing and localization. The present study investigated the relative abundance (RA) of gene transcripts in the two lineages of in vitro-produced expanded bovine blastocysts in relation to timing of development, i.e., blastocyst expansion and localization of specific mRNAs. Expanded blastocysts from either Day 7 or Day 8 or isolated ICMs derived thereof were analyzed with the aid of a semiquantitative reverse transcriptase-polymerase chain reaction assay for gene transcripts, which are thought to play a pivotal role in blastocyst expansion, i.e., Na/K-ATPase alpha1 subunit (Na/K), E-cadherin (E-cad), zonula occludens protein-1 (ZO-1), desmocollin II (Dc II), plakophilin (Plako), trophoblastic function (interferon tau [IFtau]), and glucose transport (glucose transporter-1, -3, -4 [Glut-1, -3, -4]). Total cell number, ICM cell number, or ICM/total cells ratio were similar in Day 7 and Day 8 expanded blastocysts. Significant differences were determined in the RA for Na/K, E-cad, Dc II, Plako, and ZO-1 transcripts between TE cells of expanded blastocysts derived from either Day 7 or Day 8. The RA of Dc II, Glut-1, and Glut-4 was significantly decreased in the ICM compared with the TE at Day 7. Similarly, the RA of Na/K, Dc II, Glut-1, and Glut-4 at Day 8 of development was significantly decreased in the ICM compared with the TE. Interestingly, no differences were observed when comparing ICMs originating from blastocysts expanded at either Day 7 or Day 8. Plako and IFtau transcripts were not detected in isolated ICMs, indicating that expression of these mRNAs is restricted to the TE. In contrast, similar expression patterns within the ICM and TE were determined for Na/K, E-cad, ZO-1, and Glut-3 mRNA. Dc II, Glut-1, and Glut-4 were more abundant in the TE than in ICM. Results show that expression of developmentally important genes is related to the two cell lineages in the early embryo and emphasize the critical role of a well controlled spatial gene expression pattern for regular preimplantation development.
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PMID:Timing of blastocyst expansion affects spatial messenger RNA expression patterns of genes in bovine blastocysts produced in vitro. 1260 28

In the present study we tested the hypothesis that two isoforms of the regulatory subunit B of vacuolar-type ATPase (V-ATPase) are expressed in the zebrafish Danio rerio. The complete coding sequences for both isoforms, vatB1 and vatB2, were cloned and sequenced. BLASTX analysis revealed the greatest similarity to amino acid sequences of B subunits from the European eel Anguilla anguilla and rainbow trout Oncorhynchus mykiss. The isoforms were expressed in a bacterial system and the recombinant proteins verified using isoform-specific antibodies directed against vatB isoforms of the eel. The distribution of both isoforms in zebrafish tissues was investigated using reverse transcriptase-polymerase chain reaction and western blot analysis. The results revealed that at the RNA level both isoforms were expressed in all tested organs, i.e. the gills, swimbladder, heart, kidney, liver, spleen, intestine and skeletal muscle. At the protein level, however, there were tissue-specific variations in the levels of the two vatB isoforms expressed. The highest amounts of V-ATPase were detected in total protein preparations from gill, heart and liver tissue. In liver tissue, however, the western blot analysis indicated that vatB1 was not as prominent as vatB2, and immunohistochemistry revealed that antibodies directed against vatB1 yielded a very weak staining in a number of cells, while an antibody directed against vatB1 and vatB2 yielded a strong staining in virtually every cell. Similarly, neurosecretory cells of the small intestine were stained with an antibody directed against vatB1 and vatB2, but not with an antibody specific for vatB1. Therefore we conclude that the differential expression of two isoforms of the V-ATPase subunits, which may serve different functions as in several mammalian species, may also be a common phenomenon in teleost fish.
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PMID:Expression of two isoforms of the vacuolar-type ATPase subunit B in the zebrafish Danio rerio. 1272 12

Brain expression of the multidrug resistance proteins (MRPs), a collection of membrane-associated ATP-dependent efflux transporters, is poorly understood. Although several studies have examined the expression of these proteins within the brain barriers (i.e., the blood-brain barrier and choroid plexus), little information is available with respect to brain parenchyma cells such as microglia and astrocytes. Because microglia are the primary brain cells infected by the human immunodeficiency virus type 1 (HIV-1), MRP1 expression within microglia may contribute to lower brain accumulation of anti-HIV drugs. To examine the expression pattern of MRP1 within microglia, we performed reverse transcriptase-polymerase chain reaction analysis and Western blotting on a rat brain microglia cell line MLS-9, and in primary cultures of rat microglia. Both rat MRP1 (rMPR1) mRNA and protein were expressed in the cell line, as well as the primary cultures. We then characterized rMRP1-mediated transport properties in MLS-9 cells using [3H]vincristine, a known MRP1 substrate. Vincristine accumulation by monolayers of MLS-9 cells increased significantly in the presence of several well established MRP1 inhibitors (MK571, genistein, sulfinpyrazone, probenecid, and indomethacin), protease inhibitors, or the ATPase inhibitor sodium azide. In addition, vincristine accumulation was significantly modulated by altering the intracellular concentration of the reduced form of glutathione, further suggesting the involvement of rMRP1-mediated transport. These results provide strong evidence that the MRP1 protein is both expressed and functional in microglia cells. They also suggest that brain parenchyma can act as a "second" barrier to drug permeability and regulate brain distribution/accumulation of various xenobiotics, including protease inhibitors.
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PMID:Functional expression of the multidrug resistance protein 1 in microglia. 1289 36

To ascertain some of the important biochemical and molecular events that take place during early larval development of silver sea bream (Sparus sarba), we undertook a study of changes in the morphology as well as the ontogeny of the RNA-DNA ratio, growth hormone (GH), insulin-like growth factor I (IGF-I) messenger RNA abundance, Na(+)-K(+)-ATPase subunit mRNA abundance, and Na(+)-K(+)-ATPase enzyme activity. Larvae samples were collected at 1 to 46 days posthatch (dph). At 7 dph the yolk sac was fully absorbed, and from 28 dph onward larvae underwent rapid developmental changes to the juvenile stage. The RNA-DNA ratio was highest at 1 dph, decreased to low levels between 7 and 21 dph, then increased by 28 dph, and then again by 46 dph. The ontogenetic profiles of GH, IGF-I, and Na(+)-K(+)-ATPase alpha1 and beta1 subunits were studied using reverse transcriptase polymerase chain reaction, coupled with radioisotope hybridization of immobilized DNA. Growth hormone abundance reached a constant and high level from 35 dph onward, whereas the IGF-I level reached a peak at 35 dph and then significantly decreased. Both Na(+)-K(+)-ATPase alpha1 and beta1 subunit mRNAs increased up to 35 dph, however, at 46 dph the alpha1 subunit remained high whereas the beta1 subunit decreased. Na(+)-K(+)-ATPase activity was low in 1-dph larvae but increased rapidly as development progressed. The importance of these findings is discussed within the context of larval development.
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PMID:Larval development of silver sea bream (Sparus sarba): ontogeny of RNA-DNA ratio, GH, IGF-I, and Na(+)-K(+)-ATPase. 1292 22

Commercially available human complementary DNA microarrays were used to compare differential expression in the livers of Atlantic salmon ( Salmo salar) infected with Aeromonas salmonicida and of healthy fish. Complementary DNA probes were prepared from total RNA isolated from livers of control salmon and infected salmon by reverse transcription in the presence of (33)P-dCTP and independently hybridized to human GENE-FILTERS GF211 microarrays. Of the 4131 known genes on the microarray, 241 spots gave clearly detectable signals using labeled RNA from the control salmon liver. Of these, 4 spots were consistently found to have a greater than 2-fold increase in infected salmon compared with controls when using the same pair of filters to generate hybridization data from triplicates. These up-regulated genes were ADP/ATP translocase (AAT2), Na(+)/K(+) ATPase, acyloxyacyl hydrolase (AOAH), and platelet-derived growth factor (PDFG-A). A BlastN search revealed an AAT2 homolog from Atlantic salmon, and a reverse transcriptase polymerase chain reaction assay using primers based on this sequence confirmed its up-regulation (approx. 1.8-fold) during early infection. This work demonstrates the feasibility of using human microarrays to facilitate the discovery of differentially expressed genes in Atlantic salmon, for which no homologous microarrays are available.
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PMID:Use of human cDNA microarrays for identification of differentially expressed genes in Atlantic salmon liver during Aeromonas salmonicida infection. 1450 54

Cysteinyl leukotrienes are increased during acute lung injury in animals and humans. In this study, we determined the effect of leukotriene D4 (LTD4) on the function of Na,K-ATPase in alveolar epithelial cells and on alveolar fluid clearance in rat lungs. LTD4 (1 x 10(-7) M) increased Na,K-ATPase activity at 1 and 5 minutes by 14% (p < 0.05) and 31% (p < 0.001), respectively, in A549 alveolar epithelial cells. This was accompanied by recruitment of Na,K-ATPase alpha1 subunits from intracellular compartment(s) to the basolateral plasma membrane. LTD4-induced alpha1 Na,K-ATPase membrane translocation was blocked by the dual cysteinyl LT1 (cysLT1)/ cysteinyl LT3 (cysLT3) receptor antagonist BAY-u9773, but not by the cysLT1 antagonist MK571, implicating the cysLT3 receptor. Expression of mRNA for cysLT2, but not cysLT1, was confirmed in A549 cells and rat alveolar type 2 cells by reverse transcriptase-polymerase chain reaction. Finally, compared with control, LTD4 (1 x 10(-11) M) increased alveolar fluid clearance by 41% (p < 0.001) in isolated, perfused rat lungs; this was also blocked by BAY-u9773 but not MK571. By activating alveolar epithelial Na,K-ATPase and increasing alveolar fluid reabsorption, cysteinyl leukotrienes may, in part, have a beneficial role in the acute respiratory distress syndrome.
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PMID:Leukotriene D4 activates alveolar epithelial Na,K-ATPase and increases alveolar fluid clearance. 1473 32

The entire developmental cycle of the obligate intracellular bacteria Chlamydia pneumoniae takes place within the inclusion body. As many gram negative bacteria, Chlamydia possess a type III-secretion system (TTSS), which allows them to target effector molecules into the host cell. The expression and localization of several proteins constituting the TTSS apparatus and of proteins supposed to be secreted by the TTSS have been investigated. For the TTSS-constituting proteins, we selected representatives such as YscN (ATPase), LcrE (putative "lid" of the TTSS) and LcrH1 (postulated to be a chaperone). Furthermore, we focused on the putative effector proteins IncA, IncB, IncC, Cpn0809 and Cpn1020. Expression of these proteins was detected by reverse transcriptase-PCR followed by immunoblot analysis using antisera that were generated against the corresponding recombinant proteins. Thereby, expression could be detected on the RNA and/or protein level. Intracellular localization of proteins under investigation was determined by immunofluorescence assays using the respective antisera. YscN was shown to be distributed equally throughout the inclusion body, whereas LcrE gave a more prominent staining of the inclusion membrane. IncA was detected mainly on the membrane of the inclusion body, whereas IncB and IncC were shown to be located within the inclusion. Immunofluorescence assays with antisera raised against Cpn0809 and Cpn1020 showed completely different labeling. Signals corresponding to Cpn0809 and Cpn1020 were distributed within the host cell rather than inside the inclusions. Taken together, the different localization patterns of the effector proteins indicate differences in function and interplay with the host cell.
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PMID:Expression and localization of type III secretion-related proteins of Chlamydia pneumoniae. 1459 77

Bacillus megaterium QM B1551 plasmid pBM400, one of seven indigenous plasmids, has been labeled with a selectable marker, isolated, completely sequenced, and partially characterized. A sequence of 53,903 bp was generated, revealing a total of 50 predicted open reading frames (ORFs); 33 were carried on one strand and 17 were carried on the other. These ORFs comprised 57% of the pBM400 sequence. Besides the replicon region and a complete rRNA operon that have previously been described, several interesting genes were found, including genes for predicted proteins for cell division (FtsZ and FtsK), DNA-RNA interaction (FtsK, Int/Rec, and reverse transcriptase), germination (CwlJ), styrene degradation (StyA), and heavy metal resistance (Cu-Cd export and ATPase). Three of the ORF products had high similarities to proteins from the Bacillus anthracis virulence plasmid pXO1. An insertion element with similarity to the IS256 family and several hypothetical proteins similar to those from the chromosomes of other Bacillus and Lactococcus species were present. This study provides a basis for isolation and sequencing of other high-molecular-weight plasmids from QM B1551 and for understanding the role of megaplasmids in gram-positive bacteria. The genes carried by pBM400 suggest a possible role of this plasmid in the survival of B. megaterium in hostile environments with heavy metals or styrene and also suggest that there has been an exchange of genes within the gram-positive bacteria, including pathogens.
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PMID:Sequencing and characterization of pBM400 from Bacillus megaterium QM B1551. 1460 53

Na/K-ATPase plays an important role in ion regulation in teleost fishes. In most taxa several isoforms exist to provide physiological versatility to specific cell types, but little is known about Na/K-ATPase isoforms in fish. A reverse transcriptase polymerase chain reaction approach was used to identify Na/K-ATPase a-subunit isoforms from the gill and muscle of an estuarine teleost, Fundulus heteroclitus. Full-length complementary DNA sequences were similar at both the nucleotide level (74.4%) and the amino acid level (82.5%). Phylogenetic analysis indicated that the gill isoform was similar to the mammalian a1 subunit and the muscle isoform was similar to the mammalian a2 subunit. Northern blotting and isoform-specific polymerase chain reaction were used to determine the tissue distribution of the isoforms. The gill (a1) isoform was expressed in all tissues, while the muscle (a2) isoform had a much more restricted expression pattern, being present at high levels only in muscle and brain, consistent with the distributions of these isoforms in mammals.
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PMID:Molecular cloning and characterization of two Na/K-ATPase isoforms in Fundulus heteroclitus. 1496 Dec 45

The objective of the present study was to test the hypothesis that fish gills can express more than one isoform of the Na+-K+-ATPase a subunit responsible for ion regulation in seawater and freshwater environments. Using rapid amplification of complementary DNA ends (RACE), we cloned and sequenced full-length cDNAs encoding Na+-K+-ATPase alpha 1 and alpha 3 subunits of tilapia (Oreochromis mossambicus). Clone TG33 is 3390 bp in length and encodes a polypeptide of 1023 amino acids, while clone TH3 is 3581 bp in length and encodes a protein of 1010 amino acids. Clones TG33 and TH3 showed 91% and 88% identities at the amino acid level with previously described animal Na+-K+-ATPase alpha 1 and alpha 3 subunits, respectively. Northern blot and reverse transcriptase polymerase chain reaction analyses indicated that the alpha 1 subunit is expressed predominantly in kidney and intestine, while the alpha 3 subunit is expressed mainly in brain and heart. However, lower levels of expression of both genes were detected in other tissues such as gill, spleen, and testis. The amounts of both alpha 1 and alpha 3 subunit messenger RNA in gill tissue increased with the level of environmental salinity. This provides direct evidence of enhanced transcription of N+-K+-ATPase alpha 1 and alpha 3 subunit genes upon salinity challenge.
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PMID:Gene expression of Na+-K+-ATPase alpha 1 and alpha 3 subunits in gills of the teleost Oreochromis mossambicus, adapted to different environmental salinities. 1496 Dec 49


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