JOURNAL OF PHYCOLOGY, Issue 1 2005
Misaki Takabayashi
To understand the enhanced ability of marine diatoms to assimilate nitrogen (N), we measured changes in the transcript abundance and enzyme activity of glutamine synthetase (GS), one of the key enzymes that link carbon (C) and N metabolism, in the common diatom Skeletonema costatum (Greville) Cleve. Transcript abundance of glnII (the gene that encodes the GSII isoenzyme), measured by quantitative reverse transcriptase-PCR, and total GS activity increased 2 to 3.5 times above background in the cells taking up nitrate (NO3,) but not the cells taking up ammonium (NH4+). A background level of glnII mRNA was maintained at a steady level up to 15 days of N starvation before decreasing to below detection after 21 days. These results confirm that transcription of glnII is induced to assimilate NH4+ derived from reduction of NO3,. Because of this role of GSII in diatoms assimilating NH4+ derived from NO3, reduction rather than from the environmental NH4+, quantification of glnII mRNA promises to be a useful indication of new production by phytoplankton. [source]

MODULATION OF SIGNAL TRANSDUCERS AND ACTIVATORS OF TRANSCRIPTION (STAT) FACTOR PATHWAYS DURING FOCAL CEREBRAL ISCHAEMIA: A GENE EXPRESSION ARRAY STUDY IN RAT HIPPOCAMPUS AFTER MIDDLE CEREBRAL ARTERY OCCLUSION

CLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 11 2007
Sheng-Li Sun
SUMMARY 1Signal transducers and activators of transcription (STAT) factors are a family of transcription factors that mediate intracellular signalling initiated at cytokine cell surface receptors and transmitted to the nucleus. In the present study, we determined the global changes in STAT gene expression in the hippocampus of rats after focal cerebral ischaemia and reperfusion using microarray analysis. 2The present study used middle cerebral artery occlusion (MCAO) to induce ischaemia and reperfusion in Sprague-Dawley rats. Using superarray Q series Janus tyrosine kinases (Jak)/STAT signalling pathway gene array, a total of 96 genes was screened in adult male rat hippocampus after transient focal cerebral ischaemia. 3The results showed that 23 genes were upregulated at least twofold by ischaemia treatment and that 12 genes were downregulated at least threefold by ischaemia treatment compared with controls. 4After confirmation by quantitative real-time polymerase chain reaction, the data suggest that the gene expression of STAT2, 5a, 5b, 6 and suppressor of cytokine signalling (SOCS) 4 was increased by ischaemia, probably due to a compensatory response of the brain, which may play a protective role in damaged brain tissue. 5The results of the present study provide evidence on global changes in STAT gene expression in the hippocampus of rats after focal cerebral ischaemia and reperfusion, in which STAT2, 5a, 5b, 6 and SOCS4 were confirmed to be significantly modulated during focal cerebral ischaemia. [source]

Plasmodium falciparum myosins: Transcription and translation during asexual parasite development,

CYTOSKELETON, Issue 4 2005
Jacqueline Chaparro-Olaya
Abstract Six myosins genes are now annotated in the Plasmodium falciparum Genome Project. Malaria myosins have been named alphabetically; accordingly, we refer to the two latest additions as Pfmyo-E and Pfmyo-F. Both new myosins contain regions characteristic of the functional motor domain of "true" myosins and, unusually for P. falciparum myosins, Pfmyo-F encodes two consensus IQ light chain-binding motifs. Phylogenetic analysis of the 17 currently known apicomplexan myosins together with one representative of each myosin class clusters all but one of the apicomplexan sequences together in Class XIV. This refines the earlier definition of the Class XIV Subclasses XIVa and XIVb. RT-PCR on blood stage parasite mRNA amplifies a specific product for all six myosins and each shows developmentally regulated transcription. Thus: Pfmyo-A and Pfmyo-B genes are transcribed throughout development; Pfmyo-C is predominant in trophozoites; Pfmyo-D occurs in trophozoites and schizonts; Pfmyo-E though barely present in earlier stages is abundant in schizonts; Pfmyo-F increases steadily throughout development and maturation. It is known that Pfmyo-A and Pfmyo-B are synthesised during late schizogony and we now show that Pfmyo-D expression is also temporally regulated to late trophozoites and schizonts where it distributes close to segregating nuclei. Thus, in asexual stages myosin synthesis does not always parallel transcript accumulation, showing that translation is also regulated. The implication is that the mRNAs are either subjected to turnover, synthesised and degraded, or that they are sequestered in an inactivate form until required for protein synthesis. Cell Motil. Cytoskeleton 60:200,213, 2005. © 2005 Wiley-Liss, Inc. [source]

Three types of cilia including a novel 9+4 axoneme on the notochordal plate of the rabbit embryo

DEVELOPMENTAL DYNAMICS, Issue 12 2006
Kerstin Feistel
Abstract Motile monocilia play a pivotal role in left-right axis determination in mouse and zebrafish embryos. Cilia with 9+0 axonemes localize to the distal indentation of the mouse egg cylinder ("node"), while Kupffer's vesicle cilia in zebrafish show 9+2 arrangements. Here we studied cilia in a prototype mammalian embryo, the rabbit, which develops via a flat blastodisc. Transcription of ciliary marker genes Foxj1, Rfx3, lrd, polaris, and Kif3a initiated in Hensen's node and persisted in the nascent notochord. Cilia emerged on cells leaving Hensen's node anteriorly to form the notochordal plate. Cilia lengthened to about 5 ,m and polarized from an initially central position to the posterior pole of cells. Electron-microscopic analysis revealed 9+0 and 9+2 cilia and a novel 9+4 axoneme intermingled in a salt-and-pepper-like fashion. Our data suggest that despite a highly conserved ciliogenic program, which initiates in the organizer, axonemal structures may vary widely within the vertebrates. Developmental Dynamics 235:3348,3358, 2006. © 2006 Wiley-Liss, Inc. [source]

Homologues of nitrite reductases in ammonia-oxidizing archaea: diversity and genomic context

ENVIRONMENTAL MICROBIOLOGY, Issue 4 2010
Rita Bartossek
Summary Ammonia-oxidizing archaea are frequent and ubiquitous inhabitants of terrestrial and marine environments. As they have only recently been detected, most aspects of their metabolism are yet unknown. Here we report on the occurrence of genes encoding potential homologues of copper-dependent nitrite reductases (NirK) in ammonia-oxidizing archaea of soils and other environments using metagenomic approaches and PCR amplification. Two pairs of highly overlapping 40 kb genome fragments, each containing nirK genes of archaea, were isolated from a metagenomic soil library. Between 68% and 85% of the open reading frames on these genome fragments had homologues in the genomes of the marine archaeal ammonia oxidizers Nitrosopumilus maritimus and Cenarchaeum symbiosum. Extensions of NirK homologues with C-terminal fused amicyanin domains were deduced from two of the four fosmids indicating structural variation of these multicopper proteins in archaea. Phylogenetic analyses including all major groups of currently known NirK homologues revealed that the deduced protein sequences of marine and soil archaea were separated into two highly divergent lineages that did not contain bacterial homologues. In contrast, another separated lineage contained potential multicopper oxidases of both domains, archaea and bacteria. More nirK gene variants directly amplified by PCR from several environments indicated further diversity of the gene and a widespread occurrence in archaea. Transcription of the potential archaeal nirK in soil was demonstrated at different water contents, but no significant increase in transcript copy number was observed with increased denitrifying activity. [source]

Deposition of chromatin-IgG complexes in skin of nephritic MRL-lpr/lpr mice is associated with increased local matrix metalloprotease activities

EXPERIMENTAL DERMATOLOGY, Issue 8 2010
Annica Hedberg
Please cite this paper as: Deposition of chromatin-IgG complexes in skin of nephritic MRL-lpr/lpr mice is associated with increased local matrix metalloprotease activities. Experimental Dermatology 2010; 19: e265,e274. Abstract:, Chromatin-IgG complexes appear as electron dense structures (EDS) in glomerular basement membranes in lupus nephritis. Here, we present results of comparative analyses of the composition of EDS in murine lupus dermatitis and nephritis. One focus was to perform an analytical approach to understand why such complex structures bind skin basement membrane components. Transcription of skin membrane-encoding genes was analysed to see if expression of such genes was increased, eventually indicating that binding capacity of immune complexes increased when dermatitis developed. Variations in matrix metalloprotease 2 (MMP2), MMP9 and Dnase1 mRNA levels and enzymatic activities were correlated with circulatory chromatin-IgG complexes and deposition in skin. We also examined if glomerular deposits of EDS predicted similar deposits in skin of (NZB × NZW)F1 or MRL-lpr/lpr mice, as we observed chromatin-IgG complexes in capillary lumina in skin and glomeruli in both strains. EDS consisting of chromatin fragments and IgG were found sub-epidermally in skin with LE-like lesions of end-stage nephritic MRL-lpr/lpr mice. Dermal MMP-encoding genes were up-regulated during disease progression, and gelatinolytic activity was increased in affected skin. Dnase1 mRNA level and total nuclease activity remained stable in skin during the disease, in contrast to progressive loss of renal Dnase1 mRNA and total renal nuclease activity during development of nephritis. Loss of renal Dnase1 may explain release of chromatin fragments, while increased MMP activity may disrupt membranes making them accessible for chromatin fragment-IgG complexes. Circulatory chromatin-IgG complexes, and up-regulated intradermal MMP activity may be crucial for deposition of immune complexes in skin of lupus-prone mice. [source]

The Effects of Steroid Hormones on the Transcription of Genes Encoding Enzymes of Oxidative Phosphorylation

EXPERIMENTAL PHYSIOLOGY, Issue 1 2003
Klaus Scheller
Regulation of energy metabolism is one of the major functions of steroid hormones. In this process, mitochondria, by way of oxidative phosphorylation, play a central role. Depending on the energy needs of the cell, on the tissue, on the developmental stage and on the intensity of the hormonal stimulus, the response can be an activation of pre-existing respiratory chain components, an increased transcription of nuclear-encoded and/or mitochondrial-encoded respiratory chain enzyme (OXPHOS) genes and of biosynthesis of the respective enzyme subunits or, in extreme cases of high energy needs, an increase in the number of mitochondria and mitochondrial DNA content per cell. Some of the hormonally regulated systems involving effects on nuclear and mitochondrial OXPHOS genes are reviewed in this paper. The possible molecular mechanisms of steroid hormone action on nuclear and mitochondrial gene transcription and possible ways of coordination of transcription in these two separate cell compartments involving direct interaction of steroid receptors with hormone response elements in nuclear OXPHOS genes and in mitochondria and induction/activation of nuclear-encoded regulatory factors affecting mitochondrial gene transcription are presented. [source]

Loss of sense transgene-induced post-transcriptional gene silencing by sequential introduction of the same transgene sequences in tobacco

FEBS JOURNAL, Issue 7 2010
Sayaka Hirai
RNA silencing is an epigenetic inhibition of gene expression and is guided by small interfering RNAs. Sense transgene-induced post-transcriptional gene silencing (S-PTGS) occurs in a portion of a transgenic plant population. When a sense transgene encoding a tobacco endoplasmic reticulum ,-3 fatty acid desaturase (NtFAD3) was introduced into tobacco plants, an S-PTGS line, S44, was obtained. Introduction of another copy of the NtFAD3 transgene into S44 plants caused a phenotypic change from S-PTGS to overexpression. Because this change was associated with the methylation of the promoter sequences of the transgene, reduced transcriptional activity may abolish S-PTGS and residual transcription of the sense transgene may account for the overexpression. To clarify whether RNA-directed DNA methylation (RdDM) can repress the transcriptional activity of the S44 transgene locus, we introduced several RdDM constructs targeting the transgene promoter. An RdDM construct harboring a 200-bp-long fragment of promoter sequences efficiently abrogated the generation of NtFAD3 small interfering RNAs in S44 plants. Transcription of the transgene was partially repressed, but the resulting NtFAD3 mRNAs successfully accumulated and an overexpressed phenotype was established. Our results indicate an example in which overexpression of the transgene is established by complex epigenetic interactions among the transgenic loci. [source]

Transcription of mammalian cytochrome c oxidase subunit IV-2 is controlled by a novel conserved oxygen responsive element

FEBS JOURNAL, Issue 21 2007
Maik Hüttemann
Subunit 4 of cytochrome c oxidase (CcO) is a nuclear-encoded regulatory subunit of the terminal complex of the mitochondrial electron transport chain. We have recently discovered an isoform of CcO 4 (CcO4-2) which is specific to lung and trachea, and is induced after birth. The role of CcO as the major cellular oxygen consumer, and the lung-specific expression of CcO4-2, led us to investigate CcO4-2 gene regulation. We cloned the CcO4-2 promoter regions of cow, rat and mouse and compared them with the human promoter. Promoter activity is localized within a 118-bp proximal region of the human promoter and is stimulated by hypoxia, reaching a maximum (threefold) under 4% oxygen compared with normoxia. CcO4-2 oxygen responsiveness was assigned by mutagenesis to a novel promoter element (5,-GGACGTTCCCACG-3,) that lies within a 24-bp region that is 79% conserved in all four species. This element is able to bind protein, and competition experiments revealed that, within the element, the four core bases 5,-TCNCA-3, are obligatory for transcription factor binding. CcO isolated from lung showed a 2.5-fold increased maximal turnover compared with liver CcO. We propose that CcO4-2 expression in highly oxygenated lung and trachea protects these tissues from oxidative damage by accelerating the last step in the electron transport chain, leading to a decrease in available electrons for free radical formation. [source]

Identification of ERR, as a specific partner of PGC-1, for the activation of PDK4 gene expression in muscle

FEBS JOURNAL, Issue 8 2006
Makoto Araki
Pyruvate dehydrogenase kinase 4 (PDK4) is a key regulatory enzyme involved in switching the energy source from glucose to fatty acids in response to physiological conditions. Transcription of the PDK4 gene is activated by fasting or by the administration of a PPAR, ligand in a tissue-specific manner. Here, we show that the two mechanisms are independent, and that ERR, is directly involved in PPAR,-independent transcriptional activation of the PDK4 gene with PGC-1, as a specific partner. This conclusion is based on the following evidence. First, detailed mutation analyses of the cloned PDK4 gene promoter sequence identified a possible ERR,-binding motif as the PGC-1, responsive element. Second, overexpression of ERR, by cotransfection enhanced, and the knockout of it by shRNAs diminished, PGC-1,-dependent activation. Third, specific binding of ERR, to the identified PGC-1, responsive sequence was confirmed by the electrophoresis mobility shift assay. Finally, cell-type-specific responsiveness to PGC-1, was observed and this could be explained by differences in the expression levels of ERR,, however, ectopic expression of ERR, in poorly responsive cells did not restore PGC-1, responsiveness, indicating that ERR, is necessary, but not sufficient for the response. [source]

Transcription of individual tRNAGly1 genes from within a multigene family is regulated by transcription factor TFIIIB

FEBS JOURNAL, Issue 20 2005
Akhila Parthasarthy
Members of a multigene family from the silkworm Bombyx mori have been classified based on their transcriptions in homologous nuclear extracts, into three groups of highly, moderately and poorly transcribed genes. Because all these gene copies have identical coding sequences and consequently identical promoter elements (the A and B boxes), the flanking sequences modulate their expression levels. Here we demonstrate the interaction of transcription factor TFIIIB with these genes and its role in regulating differential transcriptions. The binding of TFIIIB to the poorly transcribed gene -6,7 was less stable compared with binding of TFIIIB to the highly expressed copy, -1. The presence of a 5, upstream TATA sequence closer to the coding region in -6,7 suggested that the initial binding of TFIIIC to the A and B boxes sterically hindered anchoring of TFIIIB via direct interactions, leading to lower stability of TFIIIC,B-DNA complexes. Also, the multiple TATATAA sequences present in the flanking regions of this poorly transcribed gene successfully competed for TFIIIB reducing transcription. The transcription level could be enhanced to some extent by supplementation of TFIIIB but not by TATA box binding protein. The poor transcription of -6,7 was thus attributed both to the formation of a less stable transcription complex and the sequestration of TFIIIB. Availability of the transcription factor TFIIIB in excess could serve as a general mechanism to initiate transcription from all the individual members of the gene family as per the developmental needs within the tissue. [source]

Biochemical and molecular characterization of a laccase from the edible straw mushroom, Volvariella volvacea

FEBS JOURNAL, Issue 2 2004
Shicheng Chen
We have isolated a laccase (lac1) from culture fluid of Volvariella volvacea, grown in a defined medium containing 150 µm CuSO4, by ion-exchange and gel filtration chromatography. Lac1 has a molecular mass of 58 kDa as determined by SDS/PAGE and an isoelectric point of 3.7. Degenerate primers based on the N-terminal sequence of purified lac1 and a conserved copper-binding domain were used to generate cDNA fragments encoding a portion of the lac1 protein and RACE was used to obtain full-length cDNA clones. The cDNA of lac1 contained an ORF of 1557 bp encoding 519 amino acids. The amino acid sequence from Ala25 to Asp41 corresponded to the N-terminal sequence of the purified protein. The first 24 amino acids are presumed to be a signal peptide. The expression of lac1 is regulated at the transcription level by copper and various aromatic compounds. RT-PCR analysis of gene transcription in fungal mycelia grown on rice-straw revealed that, apart from during the early stages of substrate colonization, lac1 was expressed at every stage of the mushroom developmental cycle defined in this study, although the levels of transcription varied considerably depending upon the developmental phase. Transcription of lac1 increased sharply during the latter phase of substrate colonization and reached maximum levels during the very early stages (primordium formation, pinhead stage) of fruit body morphogenesis. Gene expression then declined to ,,20,30% of peak levels throughout the subsequent stages of sporophore development. [source]

Ambient pH controls the expression of endopolygalacturonase genes in the necrotrophic fungus Sclerotinia sclerotiorum

FEMS MICROBIOLOGY LETTERS, Issue 2 2003
Pascale Cotton
Abstract In the necrotrophic fungus Sclerotinia sclerotiorum, secretion of polygalacturonases (PGs) and decrease of the environmental pH via oxalic acid production are considered as the main pathogenicity determinants. In order to evaluate the relationship between these two aspects of the infection process, we analyzed the expression of the endoPG-encoding genes pg1,3. Transcription of pg1,3 was not carbon regulated but was strictly controlled by pH and highly favored in a narrow range of acidic pH. During plant infection, a pH gradient was established in relation to oxalic acid secretion. Transcripts of pg1,3 were localized to the zone of colonization of healthy tissues while transcripts of genes encoding other lytic enzymes were restricted to the more acidic zones of the infected tissues. Our results show that progressive acidification of the ambient medium by the fungus is a major strategy for the sequential expression of pathogenicity factors. [source]

Tuning and Transcription of the Supramolecular Organization of a Fluorescent Silsesquioxane Precursor into Silica-Based Materials through Direct Photochemical Hydrolysis,Polycondensation and Micropatterning

ADVANCED FUNCTIONAL MATERIALS, Issue 3 2009
Xavier Sallenave
Abstract A new fluorescent silsequioxane precursor with tuned optical properties and controlled aggregation properties is designed. The two cyclohexyl moieties introduced in the molecular structure allow the formation of very good quality films. The J-aggregated structure is transcribed into the solid by photoacid-catalyzed hydrolysis,polycondensation. Aggregation of the chromophores is reduced and highly fluorescent materials are obtained. The photoacid generator lies on the surface of the homogeneous layer of the sol,gel precursor. This phase separation presents several advantages, including UV protection of the chromophore and easy removal of the PAG. The first example of chemical amplification in the photolithography of the conjugated silsesquioxane precursor is demonstrated. As hydrolysis,polycondensation could be achieved in a controlled way by UV exposure, chemically amplified photolithography is achieved by irradiating a composite film (,110,nm thick) on silicon wafer by using a copper TEM grid as shadow mask. The pattern is produced uniformly on a miscroscopic scale of 3,mm, the photopatterned pixels remaining highly fluorescent. The sizes of the photolithographed pixels correspond to the sizes of the rectangular holes of the 300,×,75 mesh grid (hole: 63,<$>,<$>m,×,204,<$>,<$>m). [source]

The Helicobacter hepaticus hefA Gene is Involved in Resistance to Amoxicillin

HELICOBACTER, Issue 1 2009
Clara Belzer
Abstract Background:, Gastrointestinal infections with pathogenic Helicobacter species are commonly treated with combination therapies, which often include amoxicillin. Although this treatment is effective for eradication of Helicobacter pylori, the few existing reports are less clear about antibiotic susceptibility of other Helicobacter species. In this study we have determined the susceptibility of gastric and enterohepatic Helicobacter species to amoxicillin, and have investigated the mechanism of amoxicillin resistance in Helicobacter hepaticus. Materials and methods:, The minimal inhibitory concentration (MIC) of antimicrobial compounds was determined by E -test and agar/broth dilution assays. The hefA gene of H. hepaticus was inactivated by insertion of a chloramphenicol resistance gene. Transcription was measured by quantitative real-time polymerase chain reaction. Results:, Three gastric Helicobacter species (H. pylori, H. mustelae, and H. acinonychis) were susceptible to amoxicillin (MIC < 0.25 mg/L). In contrast, three enterohepatic Helicobacter species (H. rappini, H. bilis, and H. hepaticus) were resistant to amoxicillin (MIC of 8, 16, and 6,64 mg/L, respectively). There was no detectable ,-lactamase activity in H. hepaticus, and inhibition of ,-lactamases did not change the MIC of amoxicillin of H. hepaticus. A H. hepaticus hefA (hh0224) mutant, encoding a TolC-component of a putative efflux system, resulted in loss of amoxicillin resistance (MIC 0.25 mg/L), and also resulted in increased sensitivity to bile acids. Finally, transcription of the hefA gene was not responsive to amoxicillin, but induced by bile acids. Conclusions:, Rodents are frequently colonized by a variety of enterohepatic Helicobacter species, and this may affect their global health status and intestinal inflammatory responses. Animal facilities should have treatment strategies for Helicobacter infections, and hence resistance of enterohepatic Helicobacter species to amoxicillin should be considered when designing eradication programs. [source]

Helicobacter pylori HP1034 (ylxH) is required for motility

HELICOBACTER, Issue 5 2004
Karin Van Amsterdam
ABSTRACT Background.,Helicobacter pylori motility is essential for the colonization and persistence in the human gastric mucosa. So far, more than 50 genes have been described to play a role in flagellar biosynthesis. H. pylori YlxH (HP1034) is annotated as an ATP-binding protein. However, H. pylori YlxH shows similarity to proteins involved in the flagellar biosynthesis of other bacterial species. Moreover, H. pylori ylxH is found adjacent to genes involved in flagellar biosynthesis in the sequenced genomes of H. pylori 26695 and J99. We therefore aimed to determine the role of YlxH in H. pylori motility. Materials and methods., Motility, flagellar biosynthesis and transcriptional regulation of genes encoding flagellar proteins was compared between H. pylori 11A and a knockout of ylxH in H. pylori 11A. Results., The ylxH knockout in H. pylori 11A was nonmotile on soft agar plates, whereas H. pylori 11A was motile. Furthermore, the H. pylori 11A ylxH knockout lacked flagella, while H. pylori 11A possessed two to three flagella. Transcription of H. pylori flaG (HP0751), fliM (HP1031) and fliA (HP1032) was reduced in the H. pylori 11A ylxH¯ knockout, whereas transcription of flaA (HP0601) was not altered. However, Western blot analysis showed substantially reduced amounts of the major flagellin subunit FlaA in the H. pylori 11A ylxH knockout compared to H. pylori 11A. Conclusions.,H. pylori YlxH is essential for the assembly of flagella and hence for the motility of H. pylori. [source]

Edmund Campion: Memory and Transcription By Gerard Kilroy

HISTORY, Issue 304 2006
ANNE DILLON
No abstract is available for this article. [source]

Functional polymorphism in ALOX15 results in increased allele-specific transcription in macrophages through binding of the transcription factor SPI1 ,

HUMAN MUTATION, Issue 1 2006
Jonas Wittwer
Abstract The reticulocyte-type 15-lipoxygenase-1 (ALOX15) has antiinflammatory and inflammatory effects, and is implicated in the development of asthma, arthritis, and atherosclerosis. We screened the human ALOX15 gene for variations because genetic variability in ALOX15 may influence these diseases. We detected 11 variations, including five polymorphisms located in the ALOX15 promoter region. One of these polymorphisms, a C-to-T substitution at position c.,292, created a novel transcription factor binding site for SPI1. Transcription assays revealed that promoter variants with c.,292 T transcribe twice as efficiently as all the other promoter variants containing c.,292C. This was true in macrophages that constitutively express SPI1, but not in a lung epithelial cell line that does not express SPI1. Mutation of the core-binding site for SPI1 abolished the higher transcriptional activity, and electrophoretic mobility shift assays showed that SPI1 selectively binds to the mutant c.,292 T and c.,292C promoter. These results were corroborated in primary human macrophages, in which macrophages from heterozygous c.,292CT carriers expressed three times more ALOX15 mRNA than macrophages from homozygous c.,292CC carriers. We conclude that the c.,292 T allele in the ALOX15 promoter generates a novel binding site for the transcription factor SPI1 that results in higher transcription of the gene in macrophages. This may lead to an increase in ALOX15-mediated lipid metabolites, which play a role in inflammation. Hum Mutat 27(1), 78,87, 2006. © 2005 Wiley-Liss, Inc. [source]

Transcription of major histocompatibility complex class I (Kb) and transporter associated with antigen processing 1 and 2 genes is up-regulated with age

IMMUNOLOGY, Issue 3 2004
Alain G. Assounga
Summary The transporter associated with antigen processing 1 and 2 (TAP1 and TAP2) genes belong to the ATP-binding cassette family of transporter genes. They provide peptides necessary for the assembly of major histocompatibility complex (MHC) class I molecules by transporting these peptides into the endoplasmic reticulum. As MHC class I protein expression increases with age, we have explored the effect of age on the transcription of MHC class I genes (Kb) and TAP1 and TAP2 genes in C57BL/6 mice. Blood and spleen lymphocytes were isolated from mice aged from 3 months to over 24 months. RNA was extracted and mRNA for Kb, TAP1, TAP2 was quantified using slot-blot hybridization followed by densitometry. There was a parallel age-related increase (1·5-fold) in blood lymphocyte mRNA of these genes from 3 months to 21 months. In mice over 24 months old there was a decrease in Kb and TAP1 mRNA, but an increase in TAP2 mRNA. In spleen lymphocytes an age-related increase in all three mRNA species occurred throughout life. While MHC class I and Tap genes underwent very similar age-related changes, MHC class I mRNA was about 50 times more abundant than either TAP1 or TAP2 mRNA. [source]

The accumulation of specific mRNAs following multiple blood meals in Anopheles gambiae

INSECT MOLECULAR BIOLOGY, Issue 1 2005
X. Nirmala
Abstract One approach to genetic control of transmission of the parasites that cause human malaria is based on expressing effector genes in mosquitoes that disable the pathogens. Endogenous mosquito promoter and other cis -acting DNA sequences are needed to direct the optimal tissue-, stage- and sex-specific expression of the effector molecules. The mRNA accumulation profiles of eight different genes expressed specifically in the midgut, salivary glands or fat body tissues of the malaria vector, Anopheles gambiae, were characterized as a measure of their suitability to direct the expression of effector molecules designed to disable specific stages of the parasites. RT-PCR techniques were used to determine the abundance of the gene products and their duration following multiple blood meals. Transcription from the midgut-expressed carboxypeptidase-encoding gene, AgCP, follows a cyclical, blood-inducible expression pattern with maximum accumulation every 3 h post blood meal. Other midgut-expressed genes encoding a trypsin and chymotrypsin, Antryp2 and Anchym1, respectively, and the fat body-expressed genes, Vg1 and Cathepsin, also show a blood-inducible pattern of expression with maximum accumulation 24 h after every blood meal. Expression of the Lipophorin gene in the fat body and apyrase and D7-related genes (AgApy and D7r2) in the salivary glands is constitutive and not significantly affected by blood meals. Promoters of the midgut- and fat body-expressed genes may lead to maximum accumulation of antiparasite effector molecule transcripts after multiple blood meals. The multiple feeding behaviour of An. gambiae thus can be an advantage to express high levels of antiparasite effector molecules to counteract the parasites throughout most of adult development. [source]

Curcumin downregulates H19 gene transcription in tumor cells

JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 5 2008
Renata Novak Kujund
Abstract Curcumin (diferuloymethane), a natural compound used in traditional medicine, exerts an antiproliferative effect on various tumor cell lines by an incompletely understood mechanism. It has been shown that low doses of curcumin downregulate DNA topoisomerase II alpha (TOP2A) which is upregulated in many malignances. The activity of TOP2A is required for RNA polymerase II transcription on chromatin templates. Recently, it has been reported that CTCF, a multifunctional transcription factor, recruits the largest subunit of RNA polymerase II (LS Pol II) to its target sites genome-wide. This recruitment of LS Pol II is more pronounced in proliferating cells than in fully differentiated cells. As expression of imprinted genes is often altered in tumors, we investigated the potential effect of curcumin treatment on transcription of the imprinted H19 gene, located distally from the CTCF binding site, in human tumor cell lines HCT 116, SW 620, HeLa, Cal 27, Hep-2 and Detroit 562. Transcription of TOP2A and concomitantly H19 was supressed in all tumor cell lines tested. Monoallelic IGF2 expression was maintained in curcumin-treated cancer cells, indicating the involvement of mechanism/s other than disturbance of CTCF insulator function at the IGF2/H19 locus. Curcumin did not alter H19 gene transcription in primary cell cultures derived from normal human tissues. J. Cell. Biochem. 104: 1781,1792, 2008. © 2008 Wiley-Liss, Inc. [source]

Genome-wide Characterization of Long Terminal Repeat -retrotransposons in Apple Reveals the Differences in Heterogeneity and Copy Number between Ty1 -copia and Ty3 -gypsy Retrotransposons

JOURNAL OF INTEGRATIVE PLANT BIOLOGY, Issue 9 2008
Hai-Yue Sun
Abstract The conserved domains of reverse transcriptase (RT) genes of Ty1- copia and Ty3- gypsy groups of long terminal repeat (LTR) retrotransposons were isolated from the Malus domestica genome using degenerate oligonucleotide primers. Sequence analysis showed that 45% of Ty1- copia and 63% of Ty3- gypsy RT sequences contained premature stop codons and/or indels disrupting the reading frame. High heterogeneity among RT sequences of both Ty1- copia and Ty3- gypsy group retrotransposons was observed, but Ty3- gypsy group retrotransposons in the apple genome are less heterogeneous than Ty1- copia elements. Retrotransposon copy number was estimated by dot blot hybridizations for Ty1- copia (,5 000) and Ty3- gypsy (,26 000). All elements of the two types of LTR retrotransposons comprise approximately 38% of the M. domestica genome, with the Ty3- gypsy group contribution being higher (33.5%) than the Ty1- copia one (4.6%). Transcription was not detected by reverse transcription-polymerase chain reaction for either Ty1- copia or Ty3- gypsy retrotransposons in the leaves of plants in vitro or in leaf explants cultured on medium supplemented with high concentration benzylaminopurine. This research reveals the differences in heterogeneity and copy number between Ty1- copia and Ty3- gypsy retrotransposons in the apple genome. Ty1- copia retrotransposon has higher heterogeneity than Ty3- gypsy retrotransposon, but the latter has a higher copy number, which implies that Ty3- gypsy retrotransposons may play a more important role in the apple genome evolution. [source]

Transcription of rat TRPV1 utilizes a dual promoter system that is positively regulated by nerve growth factor

JOURNAL OF NEUROCHEMISTRY, Issue 1 2007
Qing Xue
Abstract The capsaicin receptor, also known as ,transient receptor potential vanilloid receptor subtype 1' (TRPV1, VR1), is an ion channel subunit expressed in primary afferent nociceptors, which plays a critical role in pain transduction and thermal hyperalgesia. Increases in nociceptor TRPV1 mRNA and protein are associated with tissue injury,inflammation. As little is understood about what controls TRPV1 RNA transcription in nociceptors, we functionally characterized the upstream portion of the rat TRPV1 gene. Two functional rTRPV1 promoter regions and their transcription initiation sites were identified. Although both promoter regions directed transcriptional activity in nerve growth factor (NGF) treated rat sensory neurons, the upstream Core promoter was the most active in cultures enriched in sensory neurons. Because NGF is a key modulator of inflammatory pain, we examined the effect of NGF on rTRPV1 transcription in PC12 cells. NGF positively regulated transcriptional activity of both rTRPV1 promoter regions in PC12 cells. We propose that the upstream regulatory region of the rTRPV1 gene is composed of a dual promoter system that is regulated by NGF. These findings support the hypothesis that NGF produced under conditions of tissue injury and/or inflammation directs an increase of TRPV1 expression in nociceptors in part through a transcription-dependent mechanism. [source]

Identification and Regulation of Genes from a Biocontrol Strain of Fusarium oxysporum

JOURNAL OF PHYTOPATHOLOGY, Issue 9 2007
D. R. Fravel
Abstract Differential display with three time points revealed that thiram altered expression of numerous genes in the biocontrol fungus Fusarium oxysporum CS-20. Of the 101 bands purified from the differential display gel, 86 were successfully cloned, and 64 sequenced. Based on nucleic acid sequences, homology to known products was found using BLASTn for 26 sequences and homology to hypothetical proteins was found for six sequences, also from Gibberella zeae. One band (BM1 24-1) showed homology to an ABC transporter from three different fungi. Because of its association with detoxification functions, the ABC transporter was selected for further study. Mycelia of CS-20 were exposed to 25 ,g active ingredient (a.i.) thiram in liquid culture for various times from 0 to 8 h. Quantitative real-time PCR was used to evaluate gene expression. At 30 min after treatment with thiram, the ABC transporter was upregulated 20- to 25-fold relative to the control treatment. The ABC transporter was upregulated 15-fold at 1 h after treatment and 10-fold at 2 h. At 8 h after treatment, there was no difference between treated and non-treated for expression of the ABC transporter. Transcription of the gene encoding EST BM1 24-1 is induced in response to thiram treatment and may function in providing resistance in F. oxysporum isolate CS-20 to fungicides and other toxins. Tolerance to toxins may be critical to the successful inclusion of CS-20 in disease control strategies in cropping systems. [source]

Polyphenolics Increase t-PA and u-PA Gene Transcription in Cultured Human Endothelial Cells

ALCOHOLISM, Issue 2 2001
Laila H. Abou-Agag
Background: Moderate red wine consumption has been associated with a reduced risk for coronary heart disease, and this cardioprotection may be mediated, in part, by promoting fibrinolysis. This protection may be attributed to the combined or perhaps synergistic effects of alcohol and other red wine components (i.e., polyphenolics). These studies were carried out to determine whether individual phenolics (i.e., catechin, epicatechin, quercetin, and resveratrol) affect fibrinolytic protein (tissue-type plasminogen activator [t-PA] and urokinase-type PA [u-PA]) e-pression and surface-localized fibrinolytic activity in cultured human umbilical vein endothelial cells (HUVECs). Methods: Cultured HUVECs were preincubated (1 hr, 37°C) in the absence or presence of varying concentrations of catechin, epicatechin, quercetin, and resveratrol (0.001,10 ,M) and then were washed and incubated for various times in the absence of phenolics. Secreted t-PA/u-PA antigen (24 hr, enzyme-linked immunoadsorbent assay) and mRNA [0,16 hr, reverse transcription-polymerase chain reaction(RT-PCR)] levels and fibrinolytic activity (direct activation of HUVEC-bound 125I-labeled glutamyl-plasminogen, quantitation of 125I-labeled M r 20 kDa plasmin light-chain) were measured. Transient transfections of cultured HUVECs were carried out with the pt-PA222/luc and pu-PA236/luc promoter constructs, by using lipofectamine. Results: Each of the phenolics similarly increased t-PA and u-PA antigen (2- to 3-fold) and mRNA (3- to 4-fold) levels, concomitant with an increase (2- to 3-fold) in sustained (24 hr), surface-localized fibrinolytic activity. Transcription inhibitor actinomycin D abolished the induction of t-PA and u-PA mRNA e-pression by these phenolics. Transfections with the pt-PA222/luc and pu-PA236/luc promoter constructs showed 2- to 3-fold and 2- to 4-fold increases in luciferase activity for t-PA and u-PA, respectively. Conclusions: These results demonstrate that each of these phenolics up-regulates both t-PA and u-PA gene transcription, which results in the sustained increased e-pression of surface-localized fibrinolytic activity in cultured HUVECs. Wine phenolics increase fibrinolytic activity, independent of ethanol, and it is likely that the overall cardioprotective benefits associated with moderate red wine consumption are attributable to the combined, additive, or perhaps synergistic effects of alcohol and other wine components. [source]

Clinical trial: the microbiological and immunological effects of synbiotic consumption , a randomized double-blind placebo-controlled study in active Crohn's disease

ALIMENTARY PHARMACOLOGY & THERAPEUTICS, Issue 7 2010
H. Steed
Aliment Pharmacol Ther 2010; 32: 872,883 Summary Background, Crohn's disease is an inflammatory illness in which the immune response against gut microorganisms is believed to drive an abnormal immune response. Consequently, modification of mucosal bacterial communities, and the immune effects they elicit, might be used to modify the disease state. Aim, To investigate the effects of synbiotic consumption on disease processes in patients with Crohn's disease. Methods, A randomized, double-blind placebo-controlled trial was conducted involving 35 patients with active Crohn's disease, using a synbiotic comprising Bifidobacterium longum and Synergy 1. Clinical status was scored and rectal biopsies were collected at the start, and at 3- and 6-month intervals. Transcription levels of immune markers and mucosal bacterial 16S rRNA gene copy numbers were quantified using real-time PCR. Results, Significant improvements in clinical outcomes occurred with synbiotic consumption, with reductions in both Crohn's disease activity indices (P = 0.020) and histological scores (P = 0.018). The synbiotic had little effect on mucosal IL-18, INF-, and IL-1,; however, significant reductions occurred in TNF-, expression in synbiotic patients at 3 months (P = 0.041), although not at 6 months. Mucosal bifidobacteria proliferated in synbiotic patients. Conclusion, Synbiotic consumption was effective in improving clinical symptoms in patients with active Crohn's disease. [source]

Dual effects of hepatitis C virus Core protein on the transcription of cyclin-dependent kinase inhibitor p21 gene

JOURNAL OF VIRAL HEPATITIS, Issue 4 2003
H. J. Kwun
Summary. Transcription of p21 was activated in hepatitis C virus (HCV) Core-expressing HepG2 cells where its upstream p53 was stabilized. However, this effect was not absolutely required for the activation of p21 by Core, as demonstrated in Hep3B cells. In addition, an opposite effect on the transcription of p21 was observed in NIH3T3 and primary hepatocytes, where p53 was not decreased by Core. To explain the p53-independent regulation of p21 by Core, we identified a Core-responsive element between positions ,74 and ,83 of the p21 promoter, exactly overlapped with a tumour growth factor , (TGF- ,)/butyrate responsive element. Furthermore, we demonstrated that Core could activate the p21 through the element by stimulating a butyrate pathway, whereas this was inhibited through a TGF- , pathway. The opposing effects of Core protein on the transcription of p21 might be important in understanding the progression of hepatic disease in HCV-positive patients. [source]

Review article: transcriptional events controlling the terminal differentiation of intestinal endocrine cells

ALIMENTARY PHARMACOLOGY & THERAPEUTICS, Issue 2000
H. Mutoh
Summary Secretin-producing enteroendocrine cells arise from a multipotential endocrine progenitor in the crypts of the small intestine. As these cells migrate up the crypt-villus axis, they produce secretin and stop dividing as they terminally differentiate and die. Transcription of the secretin gene is controlled by a complex enhancer binding to multiple transcription factors. The basic helix-loop-helix protein, BETA2, binds to an E box sequence and associates with the p300 coactivator to activate transcription of the secretin gene. Basic helix-loop-helix proteins appear to play a pivotal role in the control of cellular differentiation. BETA2 induces cell cycle arrest and apoptosis in addition to activating secretin gene expression. Thus BETA2 may function as a master regulatory gene to coordinate terminal differentiation of secretin cells. [source]

Melanoma development and pigment cell transformation in xiphophorus

MICROSCOPY RESEARCH AND TECHNIQUE, Issue 6 2002
Claudia Wellbrock
As early as 1927, it was recognised that hybridisation of platyfish (Xiphophorus maculatus) and swordtails (Xiphophorus helleri) results in offspring that develop tumours according to Mendelian laws. Most obviously, the primary event, namely the cell lineage-specific overexpression of a structurally altered receptor tyrosine kinase, finds its parallel in many tumours of birds and mammals. Once expressed at high levels, this receptor, the Xiphophorus melanoma inducing receptor kinase Xmrk, shows constitutive activation. By using different pathways, Xmrk induces both proliferative as well as anti-apoptotic signalling in pigment cells finally leading to cell transformation, tumour induction, and progression. Analyses of the different signalling cascades induced by the Xmrk-receptor led to the identification of the src-kinase Fyn, the MAP kinases ERK1 and ERK2, the "Signal Transducer and Activator of Transcription" STAT5, and the PI3-kinase as its major downstream substrates. This review describes some of the genetic findings, as well as the results from the recent molecular analyses of the factors involved in the initiation and manifestation of pigment cell transformation and melanoma development in Xiphophorus. Microsc. Res. Tech. 58:456,463, 2002. © 2002 Wiley-Liss, Inc. [source]

Transcriptional processing of G4 DNA

MOLECULAR CARCINOGENESIS, Issue 4 2009
Silvia Tornaletti
Abstract Genomic DNA sequences with the ability to assume non-B form secondary structures have been recently shown to be particularly susceptible to genetic instability, an early contributing factor in human disease and cancer development. Transcription appears to play a central role in formation of these structures and in promoting instability at these sites. The subpathway of nucleotide excision DNA repair, transcription-coupled DNA repair (TCR), removes transcription-arresting damage from the transcribed strands of expressed genes, but little is known about how non-canonical DNA structures are processed when encountered by the transcription machinery. If such structures arrest transcription, they may elicit "gratuitous" TCR in which the resulting reiterative and futile repair replication might generate a significant level of mutagenesis in a frequently transcribed gene because of faulty processing in the area of transcription arrest. Here we will describe our current understanding of how TCR may be elicited at non-B DNA structures and summarize recent literature describing the behavior of RNA polymerases when encountering non-canonical DNA structures, with particular emphasis on quadruplex DNA. © 2009 Wiley-Liss, Inc. [source]