Showing papers on "Transcription (biology) published in 2000"
TL;DR: The completion of the Arabidopsis thaliana genome sequence allows a comparative analysis of transcriptional regulators across the three eukaryotic kingdoms and reveals the evolutionary generation of diversity in the regulation of transcription.
Abstract: The completion of the Arabidopsis thaliana genome sequence allows a comparative analysis of transcriptional regulators across the three eukaryotic kingdoms. Arabidopsis dedicates over 5% of its genome to code for more than 1500 transcription factors, about 45% of which are from families specific to plants. Arabidopsis transcription factors that belong to families common to all eukaryotes do not share significant similarity with those of the other kingdoms beyond the conserved DNA binding domains, many of which have been arranged in combinations specific to each lineage. The genome-wide comparison reveals the evolutionary generation of diversity in the regulation of transcription.
2,582 citations
1,920 citations
TL;DR: An eight-plasmid DNA transfection system for the rescue of infectious influenza A virus from cloned cDNA facilitates the design and recovery of both recombinant and reassortant influenza A viruses, and may also be applicable to the recovery of other RNA viruses entirely from cloning cDNA.
Abstract: We have developed an eight-plasmid DNA transfection system for the rescue of infectious influenza A virus from cloned cDNA. In this plasmid-based expression system, viral cDNA is inserted between the RNA polymerase I (pol I) promoter and terminator sequences. This entire pol I transcription unit is flanked by an RNA polymerase II (pol II) promoter and a polyadenylation site. The orientation of the two transcription units allows the synthesis of negative-sense viral RNA and positive-sense mRNA from one viral cDNA template. This pol I–pol II system starts with the initiation of transcription of the two cellular RNA polymerase enzymes from their own promoters, presumably in different compartments of the nucleus. The interaction of all molecules derived from the cellular and viral transcription and translation machinery results in the generation of infectious influenza A virus. The utility of this system is proved by the recovery of the two influenza A viruses: A/WSN/33 (H1N1) and A/Teal/HK/W312/97 (H6N1). Seventy-two hours after the transfection of eight expression plasmids into cocultured 293T and MDCK cells, the virus yield in the supernatant of the transfected cells was between 2 × 105 and 2 × 107 infectious viruses per milliliter. We also used this eight-plasmid system for the generation of single and quadruple reassortant viruses between A/Teal/HK/W312/97 (H6N1) and A/WSN/33 (H1N1). Because the pol I–pol II system facilitates the design and recovery of both recombinant and reassortant influenza A viruses, it may also be applicable to the recovery of other RNA viruses entirely from cloned cDNA.
1,522 citations
TL;DR: It is shown that gene constructs encoding intron-spliced RNA with a hairpin structure can induce post-transcriptional gene silencing with almost 100% efficiency when directed against viruses or endogenous genes.
Abstract: Post-transcriptional gene silencing (PTGS), a sequence-specific RNA degradation mechanism inherent in many life-forms, can be induced in plants by transforming them with either antisense1 or co-suppression2 constructs, but typically this results in only a small proportion of silenced individuals. Here we show that gene constructs encoding intron-spliced RNA with a hairpin structure can induce PTGS with almost 100% efficiency when directed against viruses or endogenous genes. These constructs could prove valuable in reverse genetics, genomics, engineering of metabolic pathways and protection against pathogens.
1,123 citations
TL;DR: The possibility that expanded repeat htt causes aberrant transcriptional regulation through its interaction with cellular transcription factors which may result in neuronal dysfunction and cell death in HD is raised.
Abstract: Huntington's Disease (HD) is caused by an expansion of a polyglutamine tract within the huntingtin (htt) protein. Pathogenesis in HD appears to include the cytoplasmic cleavage of htt and release of an amino-terminal fragment capable of nuclear localization. We have investigated potential consequences to nuclear function of a pathogenic amino-terminal region of htt (httex1p) including aggregation, protein-protein interactions, and transcription. httex1p was found to coaggregate with p53 in inclusions generated in cell culture and to interact with p53 in vitro and in cell culture. Expanded httex1p represses transcription of the p53-regulated promoters, p21(WAF1/CIP1) and MDR-1. httex1p was also found to interact in vitro with CREB-binding protein (CBP) and mSin3a, and CBP to localize to neuronal intranuclear inclusions in a transgenic mouse model of HD. These results raise the possibility that expanded repeat htt causes aberrant transcriptional regulation through its interaction with cellular transcription factors which may result in neuronal dysfunction and cell death in HD.
1,063 citations
TL;DR: It is hypothesized that the continuous generation of variation in the rDNA may also play a role in how species interactions develop in ecosystems under different conditions of energy input and nutrient supply.
Abstract: Ecological stoichiometry is the study of the balance of multiple chemical elements in ecological interactions. This paper reviews recent findings in this area and seeks to broaden the stoichiometric concept for use in evolutionary studies, in integrating ecological dynamics with cellular and genetic mechanisms, and in developing a unified means for studying diverse organisms in diverse habitats. This broader approach would then be considered “biological stoichiometry”. Evidence supporting a hypothesised connection between the C:N:P stoichiometry of an organism and its growth rate (the “growth rate hypothesis”) is reviewed. Various data indicate that rapidly growing organisms commonly have low biomass C:P and N:P ratios. Evidence is then discussed suggesting that low C:P and N:P ratios in rapidly growing organisms reflect increased allocation to P-rich ribosomal RNA (rRNA), as rapid protein synthesis by ribosomes is required to support fast growth. Indeed, diverse organisms (bacteria, copepods, fishes, others) exhibit increased RNA levels when growing actively. This implies that evolutionary processes that generate, directly or indirectly, variation in a major life history trait (specific growth rate) have consequences for ecological dynamics due to their effects on organismal elemental composition. Genetic mechanisms by which organisms generate high RNA, high growth rate phenotypes are discussed next, focusing on the structure and organisation of the ribosomal RNA genes (the “rDNA”). In particular, published studies of a variety of taxa suggest an association between growth rate and variation in the length and content of the intergenic spacer (IGS) region of the rDNA tandem repeat unit. In particular, under conditions favouring increased growth or yield, the number of repeat units (“enhancers”) increases (and the IGS increases in length), and transcription rates of rRNA increase. In addition, there is evidence in the literature that increased numbers of copies of rDNA genes are associated with increased growth and production. Thus, a combination of genetic mechanisms may be responsible for establishing the growth potential, and thus the RNA allocation and C:N:P composition, of an organism. Furthermore, various processes, during both sexual and asexual reproduction, can generate variation in the rDNA to provide the raw material for selection and to generate ecologically significant variation in C:N:P stoichiometry. This leads us to hypothesize that the continuous generation of such variation may also play a role in how species interactions develop in ecosystems under different conditions of energy input and nutrient supply.
1,037 citations
TL;DR: AtERFs are factors that respond to extracellular signals to modulate GCC box–mediated gene expression positively or negatively, and are concluded that AtERF genes were differentially regulated by ethylene and by abiotic stress conditions.
Abstract: Ethylene-responsive element binding factors (ERFs) are members of a novel family of transcription factors that are specific to plants. A highly conserved DNA binding domain known as the ERF domain is the unique feature of this protein family. To characterize in detail this family of transcription factors, we isolated Arabidopsis cDNAs encoding five different ERF proteins (AtERF1 to AtERF5) and analyzed their structure, DNA binding preference, transactivation ability, and mRNA expression profiles. The isolated AtERFs were placed into three classes based on amino acid identity within the ERF domain, although all five displayed GCC box–specific binding activity. AtERF1, AtERF2, and AtERF5 functioned as activators of GCC box–dependent transcription in Arabidopsis leaves. By contrast, AtERF3 and AtERF4 acted as repressors that downregulated not only basal transcription levels of a reporter gene but also the transactivation activity of other transcription factors. The AtERF genes were differentially regulated by ethylene and by abiotic stress conditions, such as wounding, cold, high salinity, or drought, via ETHYLENE-INSENSITIVE2 (EIN2)–dependent or –independent pathways. Cycloheximide, a protein synthesis inhibitor, also induced marked accumulation of AtERF mRNAs. Thus, we conclude that AtERFs are factors that respond to extracellular signals to modulate GCC box–mediated gene expression positively or negatively.
1,030 citations
TL;DR: It is shown here that transcriptional gene silencing accompanied by de novo methylation of a target promoter in plants can be triggered by a double‐stranded RNA containing promoter sequences.
Abstract: Double‐stranded RNA induces a post‐transcriptional gene silencing process, termed RNAi, in diverse organisms. It is shown here that transcriptional gene silencing accompanied by de novo methylation of a target promoter in plants can be triggered by a double‐stranded RNA containing promoter sequences. Similar to the double‐stranded RNA involved in RNAi, this promoter double‐stranded RNA, which is synthesized in the nucleus, is partially cleaved into small RNAs ∼23 nucleotides in length. Both transcriptional and post‐transcriptional gene silencing can thus be initiated by double‐stranded RNAs that enter the same degradation pathway. The results also implicate double‐stranded RNA in directing DNA methylation. Different constructs designed to produce double‐stranded promoter RNA in various ways were evaluated for their ability to induce gene silencing in tobacco and Arabidopsis . RNA hairpins transcribed from inverted DNA repeats were the most effective trans ‐acting silencing signals. This strategy could be useful for transcriptionally downregulating genes in a variety of plants.
968 citations
TL;DR: During the past decade, physical techniques such as optical tweezers and atomic force microscopy were used to study the mechanical properties of DNA at the single-molecule level and knowledge of DNA's stretching and twisting properties now permits these single- molecule techniques to be used in the study of biological processes such as DNA replication and transcription.
839 citations
TL;DR: These studies illustrate the necessity of conditional gene-targeting approaches in the mutagenesis and study of essential sex-linked genes, and indicate that OGT participation in intracellular glycosylation is essential for embryonic stem cell viability and for mouse ontogeny.
Abstract: Nuclear and cytoplasmic protein glycosylation is a widespread and reversible posttranslational modification in eukaryotic cells. Intracellular glycosylation by the addition of N-acetylglucosamine (GlcNAc) to serine and threonine is catalyzed by the O-GlcNAc transferase (OGT). This “O-GlcNAcylation” of intracellular proteins can occur on phosphorylation sites, and has been implicated in controlling gene transcription, neurofilament assembly, and the emergence of diabetes and neurologic disease. To study OGT function in vivo, we have used gene-targeting approaches in male embryonic stem cells. We find that OGT mutagenesis requires a strategy that retains an intact OGT gene as accomplished by using Cre-loxP recombination, because a deletion in the OGT gene results in loss of embryonic stem cell viability. A single copy of the OGT gene is present in the male genome and resides on the X chromosome near the centromere in region D in the mouse spanning markers DxMit41 and DxMit95, and in humans at Xq13, a region associated with neurologic disease. OGT RNA expression in mice is comparably high among most cell types, with lower levels in the pancreas. Segregation of OGT alleles in the mouse germ line with ZP3-Cre recombination in oocytes reveals that intact OGT alleles are required for completion of embryogenesis. These studies illustrate the necessity of conditional gene-targeting approaches in the mutagenesis and study of essential sex-linked genes, and indicate that OGT participation in intracellular glycosylation is essential for embryonic stem cell viability and for mouse ontogeny.
722 citations
TL;DR: It is concluded that activation of p73 provides a means for E2F-1 to induce death in the absence of p53, and the transcription of the p53 homologue p73 is induced.
Abstract: The transcription factor E2F-1 induces both cell-cycle progression and, in certain settings, apoptosis. E2F-1 uses both p53-dependent and p53-independent pathways to kill cells1,2,3,4,5,6,7,8. The p53-dependent pathway involves the induction by E2F-1 of the human tumour-suppressor protein p14ARF, which neutralizes HDM2 (human homologue of MDM2) and thereby stabilizes the p53 protein9. Here we show that E2F-1 induces the transcription of the p53 homologue p73. Disruption of p73 function inhibited E2F-1-induced apoptosis in p53-defective tumour cells and in p53-/- mouse embryo fibroblasts. We conclude that activation of p73 provides a means for E2F-1 to induce death in the absence of p53.
TL;DR: The structure of a highly active human CpG motif is defined and its molecular mechanism of action in primary human B cells is characterized.
Abstract: The vertebrate immune system recognizes bacterial DNA based on the presence of unmethylated CpG-dinucleotides in particular base contexts ("CpG motifs"). In contrast to mice, knowledge about CpG-mediated effects on human B cells is poor. In the present study we identify and determine an optimal human CpG motif. A phosphodiester oligonucleotide containing this motif strongly stimulated CD86, CD40, CD54, and MHC class II expression, IL-6 synthesis, and proliferation of primary human B cells. These effects required internalization of the oligonucleotide and endosomal maturation. The molecular mechanism of action of this CpG motif was associated with the sustained induction of the NF-kappaB p50/p65 heterodimer and of the transcription-factor complex AP-1. Transcription-factor activation by CpG DNA was preceded by increased phosphorylation of the stress kinases c-Jun N-terminal kinase and p38, and of activating transcription factor-2. In contrast to CpG, signaling through the B cell receptor led to activation of extracellular receptor kinase and to phosphorylation of a different isoform of c-Jun N-terminal kinase. These studies define the structure of a highly active human CpG motif and characterize its molecular mechanism of action in primary human B cells.
TL;DR: The results of this study demonstrate that the nature of the p53 response in diverse mRNA species depends on the levels of p53 protein in a cell, the type of inducing agent or event, and the cell type employed.
Abstract: Oligonucleotide microarrays were employed to quantitate mRNA levels from a large number of genes regulated by the p53 transcription factor. Responses to DNA damage and to zinc-inducible p53 were compared for their transcription patterns in cell culture. A cluster analysis of these data demonstrates that genes induced by gamma radiation, UV radiation, and the zinc-induced p53 form distinct sets and subsets with a few genes in common to all these treatments. Cell type- or cell line-specific p53 responses were detected. When p53 proteins were induced with zinc, the kinetics of induction or repression of mRNAs from p53-responsive genes fell into eight distinct classes, five different kinetics of induction, and three different kinetics of repression. In addition, low levels of p53 in a cell induced or repressed only a subset of genes observed at higher p53 levels. The results of this study demonstrate that the nature of the p53 response in diverse mRNA species depends on the levels of p53 protein in a cell, the type of inducing agent or event, and the cell type employed. Of 6000 genes examined for p53 regulatory responses, 107 induced and 54 repressed genes fell into categories of apoptosis and growth arrest, cytoskeletal functions, growth factors and their inhibitors, extracellular matrix, and adhesion genes.
TL;DR: The promoter recognition specificity of Escherichia coli RNA polymerase is modulated by replacement of the sigma subunit in the first step and by interaction with transcription factors in the second step, and the prediction of the expression hierarchy of approximately 4000 genes on the E. coli genome can be estimated.
Abstract: ▪ Abstract The promoter recognition specificity of Escherichia coli RNA polymerase is modulated by replacement of the σ subunit in the first step and by interaction with transcription factors in the second step. The overall differentiated state of ∼2000 molecules of the RNA polymerase in a single cell can be estimated after measurement of both the intracellular concentrations and the RNA polymerase-binding affinities for all seven species of the σ subunit and 100–150 transcription factors. The anticipated impact from this line of systematic approach is that the prediction of the expression hierarchy of ∼4000 genes on the E. coli genome can be estimated.
TL;DR: It is shown that the GR zinc binding region (ZBR), which includes the DNA binding and dimerization functions of the receptor, binds directly to theDimerization domain of the RelA subunit of NFkappaB in vitro and that the ZBR is sufficient to associate with RelA bound at NFKappaB response elements in vivo.
Abstract: Glucocorticoids repress NFκB-mediated activation of proinflammatory genes such as interleukin-8 (IL-8) and ICAM-1. Our experiments suggest that the glucocorticoid receptor (GR) confers this effect by associating through protein–protein interactions with NFκB bound at each of these genes. That is, we show that the GR zinc binding region (ZBR), which includes the DNA binding and dimerization functions of the receptor, binds directly to the dimerization domain of the RelA subunit of NFκB in vitro and that the ZBR is sufficient to associate with RelA bound at NFκB response elements in vivo. Moreover, we demonstrate in vivo and in vitro that GR does not disrupt DNA binding by NFκB. In transient transfections, we found that the GR ligand binding domain is essential for repression of NFκB but not for association with it and that GR can repress an NFκB derivative bearing a heterologous activation domain. We used chromatin immunoprecipitation assays in untransfected A549 cells to infer the mechanism by which the tethered GR represses NFκB-activated transcription. As expected, we found that the inflammatory signal TNFα stimulated preinitiation complex (PIC) assembly at the IL-8 and ICAM-1 promoters and that the largest subunit of RNA polymerase II (pol II) in those complexes became phosphorylated at serines 2 and 5 in its carboxy-terminal domain (CTD) heptapeptide repeats (YSPTSPS); these modifications are required for transcription initiation. Remarkably, GR did not inhibit PIC assembly under repressing conditions, but rather interfered with phosphorylation of serine 2 of the pol II CTD.
TL;DR: The expression of ARR5 in the apical meristems was confirmed by whole mount in situ analysis of seedlings and is consistent with a role for cytokinin in regulating cell division in vivo, and is due, at least in part, to increased transcription.
Abstract: We examined the expression of a family of Arabidopsis response regulators (ARR) and found that the steady-state levels of RNA for most are elevated very rapidly by cytokinin. Using nuclear run-on assays we demonstrated that this increase in ARR transcript levels in response to cytokinin is due, at least in part, to increased transcription. The start site of transcription for the ARR5 gene was identified using primer extension analysis. A DNA fragment comprised of 1.6 kb upstream of the ARR5 transcript start site conferred cytokinin-inducible gene expression when fused to a beta-glucuronidase reporter, confirming that the transcription rate of ARR5 is elevated by cytokinin. This reporter construct was also used to examine the spatial pattern of ARR5 expression. The highest levels of expression were observed in the root and shoot apical meristems, at the junction of the pedicle and the silique, and in the central portion of mature roots. The expression of ARR5 in the apical meristems was confirmed by whole mount in situ analysis of seedlings and is consistent with a role for cytokinin in regulating cell division in vivo.
TL;DR: This study is the first demonstration that Cbfa1 is controlled by MAPKs and suggests that this pathway has an important role in the control of osteoblast-specific gene expression.
TL;DR: Full-genome evidence that bacterial cells use discrete transcription patterns to control cell cycle progression is presented, concluding that in bacteria, as in yeast, genes involved in a given cell function are activated at the time of execution of that function.
Abstract: This report presents full-genome evidence that bacterial cells use discrete transcription patterns to control cell cycle progression. Global transcription analysis of synchronized Caulobacter crescentus cells was used to identify 553 genes (19% of the genome) whose messenger RNA levels varied as a function of the cell cycle. We conclude that in bacteria, as in yeast, (i) genes involved in a given cell function are activated at the time of execution of that function, (ii) genes encoding proteins that function in complexes are coexpressed, and (iii) temporal cascades of gene expression control multiprotein structure biogenesis. A single regulatory factor, the CtrA member of the two-component signal transduction family, is directly or indirectly involved in the control of 26% of the cell cycle-regulated genes.
TL;DR: Using fibroblast lineages in different stages of transformation, it is found that c-Myc and Sp1 were induced to a dramatic extent when cells overcame replicative senescence and obtained immortal characteristics, in association with telomerase activation.
Abstract: Telomerase activation is thought to be a critical step in cellular immortalization and carcinogenesis. The human telomerase catalytic subunit (hTERT) is a rate limiting determinant of the enzymatic activity of human telomerase. In the previous study, we identified the proximal 181 bp core promoter responsible for transcriptional activity of the hTERT gene. To identify the regulatory factors of transcription, transient expression assays were performed using hTERT promoter reporter plasmids. Serial deletion assays of the core promoter revealed that the 5′-region containing the E-box, which binds Myc/Max, as well as the 3′-region containing the GC-box, which binds Sp1, are essential for transactivation. The mutations introduced in the E-box or GC-box significantly decreased transcriptional activity of the promoter. Overexpression of Myc/Max or Sp1 led to significant activation of transcription in a cell type-specific manner, while Mad/Max introduction repressed it. However, the effects of Myc/Max on transactivation were marginal when Sp1 sites were mutated. Western blot analysis using various cell lines revealed a positive correlation between c-Myc and Sp1 expression and transcriptional activity of hTERT. Using fibroblast lineages in different stages of transformation, we found that c-Myc and Sp1 were induced to a dramatic extent when cells overcame replicative senescence and obtained immortal characteristics, in association with telomerase activation. These findings suggest that c-Myc and Sp1 cooperatively function as the major determinants of hTERT expression, and that the switching functions of Myc/Max and Mad/Max might also play roles in telomerase regulation.
TL;DR: It is indicated that mitochondrial ROS are required for the hypoxic activation of NF-κB and TNF-α gene transcription, but not for the LPS activation of FiB, indicating that LPS activates NF-β and TNI gene transcription through a ROS-independent mechanism.
Abstract: The transcription factor NF-kappa B stimulates the transcription of proinflammatory cytokines including TNF-alpha. LPS (endotoxin) and hypoxia both induce NF-kappa B activation and TNF-alpha gene transcription. Furthermore, hypoxia augments LPS induction of TNF-alpha mRNA. Previous reports have indicated that antioxidants abolish NF-kappa B activation in response to LPS or hypoxia, which suggests that reactive oxygen species (ROS) are involved in NF-kappa B activation. This study tested whether mitochondrial ROS are required for both NF-kappaB activation and the increase in TNF-alpha mRNA levels during hypoxia and LPS. Our results indicate that hypoxia (1.5% O2) stimulates NF-kappa B and TNF-alpha gene transcription and increases ROS generation as measured by the oxidant sensitive dye 2',7'-dichlorofluorescein diacetate in murine macrophage J774.1 cells. The antioxidants N-acetylcysteine and pyrrolidinedithiocarbamic acid abolished the hypoxic activation of NF-kappa B, TNF-alpha gene transcription, and increases in ROS levels. Rotenone, an inhibitor of mitochondrial complex I, abolished the increase in ROS signal, the activation of NF-kappa B, and TNF-alpha gene transcription during hypoxia. LPS stimulated NF-kappa B and TNF-alpha gene transcription but not ROS generation in J774.1 cells. Rotenone, pyrrolidinedithiocarbamic acid, and N-acetylcysteine had no effect on the LPS stimulation of NF-kappa B and TNF-alpha gene transcription, indicating that LPS activates NF-kappa B and TNF-alpha gene transcription through a ROS-independent mechanism. These results indicate that mitochondrial ROS are required for the hypoxic activation of NF-kappa B and TNF-alpha gene transcription, but not for the LPS activation of NF-kappa B.
TL;DR: It is shown that p53 messenger RNA levels are low in a large proportion of breast tumours and loss of expression of p53 in human breast cancer may be primarily due to lack ofexpression of HOXA5.
Abstract: Expression of the p53 gene protects cells against malignant transformation1,2. Whereas control of p53 degradation has been a subject of intense scrutiny, little is known about the factors that regulate p53 synthesis1,2. Here we show that p53 messenger RNA levels are low in a large proportion of breast tumours. Seeking potential regulators of p53 transcription, we found consensus HOX binding sites3,4 in the p53 promoter5. Transient transfection of Hox/HOXA5 activated the p53 promoter. Expression of HOXA5 in epithelial cancer cells expressing wild-type p53, but not in isogenic variants lacking the p53 gene6, led to apoptotic cell death. Moreover, breast cancer cell lines and patient tumours display a coordinate loss of p53 and HOXA5 mRNA and protein expression. The HOXA5 promoter region was methylated in 16 out of 20 p53-negative breast tumour specimens. We conclude that loss of expression of p53 in human breast cancer may be primarily due to lack of expression of HOXA5.
TL;DR: Flavopiridol found that the flavonoid potently inhibited transcription by RNA polymerase II in vitro by blocking the transition into productive elongation, a step controlled by P-TEFb.
TL;DR: Results indicate that reaper is a direct transcriptional target of Drosophila p53 following DNA damage, and a multimer of this site is sufficient for radiation induction in vivo.
TL;DR: 6S RNA represses expression from a sigma70-dependent promoter during stationary phase and suggests that the interaction of 6S RNA with RNA polymerase modulates s Sigma70-holoenzyme activity.
TL;DR: It remains entirely unclear, however, if oxidation or nitrosylation of eukaryotic transcription factors is an important mode of regulation, or whether transcriptional activating pathways are principally controlled at other redox‐sensitive levels.
Abstract: A growing body of evidence suggests that the cellular response to oxidative and nitrosative stress is primarily regulated at the level of transcription. Posttranslational modification of transcription factors may provide a mechanism by which cells sense these redox changes. In bacteria, for example, OxyR senses redox-related changes via oxidation or nitrosylation of a free thiol in the DNA binding region. This mode of regulation may serve as a paradigm for redox-sensing by eukaryotic transcription factors as most-including NF-kappaB, AP-1, and p53-contain reactive thiols in their DNA binding regions, the modification of which alters binding in vitro. Several of these transcription factors have been found to be sensitive to both reactive oxygen species and nitric oxide-related species in vivo. It remains entirely unclear, however, if oxidation or nitrosylation of eukaryotic transcription factors is an important mode of regulation, or whether transcriptional activating pathways are principally controlled at other redox-sensitive levels.-Marshall, H. E., Merchant, K., Stamler, J. S. Nitrosation and oxidation in the regulation of gene expression.
TL;DR: The isolation of an1 by transposon tagging is reported, providing genetic evidence that the transcription factors encoded by an1, an2, and an4 operate in an unexpectedly complex regulatory hierarchy.
Abstract: The petunia loci anthocyanin1 (an1), an2, an4, and an11 are required for the transcription of anthocyanin biosynthetic genes in floral organs. The an2 and an11 loci were recently cloned and shown to encode a MYB-domain transcriptional activator and a cytosolic WD40 protein, respectively. Here, we report the isolation of an1 by transposon tagging. an1 encodes a new member of the basic helix-loop-helix family of transcription factors that is functionally and evolutionarily distinct from JAF13, the apparent petunia ortholog of maize RED1 and snapdragon DELILA. We provide genetic evidence that the transcription factors encoded by an1, an2, and an4 operate in an unexpectedly complex regulatory hierarchy. In leaves, ectopic expression of AN2 induces an1 expression, whereas in anthers, an1 expression depends on an4, encoding (or controlling) a MYB protein that is paralogous to AN2. Experiments with transgenic plants expressing a post-translationally controlled AN1-GLUCOCORTICOID RECEPTOR fusion protein indicated that independent of protein synthesis, AN1 directly activates the expression of the dfrA gene encoding the enzyme dihydroflavonol 4-reductase and of Pmyb27 encoding a MYB-domain protein of unknown function.
TL;DR: The path of the nucleic acids through a transcription elongation complex was tracked by mapping cross-links between bacterial RNA polymerase and transcript RNA or template DNA onto the x-ray crystal structure and the resulting model provides insight into the functional properties of the transcription complex.
Abstract: The path of the nucleic acids through a transcription elongation complex was tracked by mapping cross-links between bacterial RNA polymerase (RNAP) and transcript RNA or template DNA onto the x-ray crystal structure. In the resulting model, the downstream duplex DNA is nestled in a trough formed by the β′ subunit and enclosed on top by the β subunit. In the RNAP channel, the RNA/DNA hybrid extends from the enzyme active site, along a region of the β subunit harboring rifampicin resistance mutations, to the β′ subunit “rudder.” The single-stranded RNA is then extruded through another channel formed by the β-subunit flap domain. The model provides insight into the functional properties of the transcription complex.
TL;DR: A high-resolution “genome array” for the study of gene expression and regulation in Escherichia coli detected transcripts for 97% and 87% of the ORFs in stationary and log phases, and transcription of between 3,000 and 4,000 predicted ORFs was observed from the antisense strand, indicating that most of the genome is transcribed at a detectable level.
Abstract: We have developed a high-resolution “genome array” for the study of gene expression and regulation in Escherichia coli. This array contains on average one 25-mer oligonucleotide probe per 30 base pairs over the entire genome, with one every 6 bases for the intergenic regions and every 60 bases for the 4,290 open reading frames (ORFs). Twofold concentration differences can be detected at levels as low as 0.2 messenger RNA (mRNA) copies per cell, and differences can be seen over a dynamic range of three orders of magnitude. In rich medium we detected transcripts for 97% and 87% of the ORFs in stationary and log phases, respectively. We found that 1,529 transcripts were differentially expressed under these conditions. As expected, genes involved in translation were expressed at higher levels in log phase, whereas many genes known to be involved in the starvation response were expressed at higher levels in stationary phase. Many previously unrecognized growth phase-regulated genes were identified, such as a putative receptor (b0836) and a 30S ribosomal protein subunit (S22), both of which are highly upregulated in stationary phase. Transcription of between 3,000 and 4,000 predicted ORFs was observed from the antisense strand, indicating that most of the genome is transcribed at a detectable level. Examples are also presented for high-resolution array analysis of transcript start and stop sites and RNA secondary structure.
TL;DR: It is demonstrated here that mutations in the serine- and arginine-rich domain and RNA recognition motif of P GC-1 interfere with the ability of PGC-1 to induce mRNAs of target genes and disrupt the ability to co-localize and associate with RNA processing factors.
Patent•
02 Oct 2000
TL;DR: In this paper, the use of expression vectors harboring the coding sequence of a gene of interest under the transcriptional control of promoting sequences for which activity is regulated by the presence of nitrogen.
Abstract: The present invention relates to a method of regulating the transcription of transgene in genetically-modified organisms. More specifically, the invention relates to the use of expression vectors harboring the coding sequence of a gene of interest under the transcriptional control of promoting sequences for which activity is regulated by the presence of nitrogen.