相关论文
2004 - Development
Mili, a mammalian member of piwi family gene, is essential for spermatogenesis

The piwi family genes, which are defined by conserved PAZ and Piwi domains, play important roles in stem cell self-renewal, RNA silencing, and translational regulation in various organisms. To reveal the function of the mammalian homolog of piwi, we produced and analyzed mice with targeted mutations in the Mili gene, which is one of three mouse homologs of piwi. Spermatogenesis in the MILI-null mice was blocked completely at the early prophase of the first meiosis, from the zygotene to early pachytene, and the mice were sterile. However, primordial germ cell development and female germ cell production were not disturbed. Furthermore, MILI bound to MVH, which is an essential factor during the early spermatocyte stage. The similarities in the phenotypes of the MILI- and MVH-deficient mice and in the physical binding properties of MILI and MVH indicate a functional association of these proteins in post-transcriptional regulation. These data indicate that MILI is essential for the differentiation of spermatocytes.

2005 - The Journal of experimental medicine
Disruption of Nrf2 enhances susceptibility to severe airway inflammation and asthma in mice

Oxidative stress has been postulated to play an important role in the pathogenesis of asthma; although a defect in antioxidant responses has been speculated to exacerbate asthma severity, this has been difficult to demonstrate with certainty. Nuclear erythroid 2 p45-related factor 2 (Nrf2) is a redox-sensitive basic leucine zipper transcription factor that is involved in the transcriptional regulation of many antioxidant genes. We show that disruption of the Nrf2 gene leads to severe allergen-driven airway inflammation and hyperresponsiveness in mice. Enhanced asthmatic response as a result of ovalbumin sensitization and challenge in Nrf2-disrupted mice was associated with more pronounced mucus cell hyperplasia and infiltration of eosinophils into the lungs than seen in wild-type littermates. Nrf2 disruption resulted in an increased expression of the T helper type 2 cytokines interleukin (IL)-4 and IL-13 in bronchoalveolar lavage fluid and in splenocytes after allergen challenge. The enhanced severity of the asthmatic response from disruption of the Nrf2 pathway was a result of a lowered antioxidant status of the lungs caused by lower basal expression, as well as marked attenuation, of the transcriptional induction of multiple antioxidant genes. Our studies suggest that the responsiveness of Nrf2-directed antioxidant pathways may act as a major determinant of susceptibility to allergen-mediated asthma.

2003 - Proceedings of the National Academy of Sciences of the United States of America
Parallel changes in gene expression after 20,000 generations of evolution in Escherichia coli

Twelve populations of Escherichia coli, derived from a common ancestor, evolved in a glucose-limited medium for 20,000 generations. Here we use DNA expression arrays to examine whether gene-expression profiles in two populations evolved in parallel, which would indicate adaptation, and to gain insight into the mechanisms underlying their adaptation. We compared the expression profile of the ancestor to that of clones sampled from both populations after 20,000 generations. The expression of 59 genes had changed significantly in both populations. Remarkably, all 59 were changed in the same direction relative to the ancestor. Many of these genes were members of the cAMP-cAMP receptor protein (CRP) and guanosine tetraphosphate (ppGpp) regulons. Sequencing of several genes controlling the effectors of these regulons found a nonsynonymous mutation in spoT in one population. Moving this mutation into the ancestral background showed that it increased fitness and produced many of the expression changes manifest after 20,000 generations. The same mutation had no effect on fitness when introduced into the other evolved population, indicating that a mutation of similar effect was present already. Our study demonstrates the utility of expression arrays for addressing evolutionary issues including the quantitative measurement of parallel evolution in independent lineages and the identification of beneficial mutations.

2010 - FEMS microbiology ecology
Archaea rather than bacteria control nitrification in two agricultural acidic soils.

Nitrification is a central component of the global nitrogen cycle. Ammonia oxidation, the first step of nitrification, is performed in terrestrial ecosystems by both ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA). Published studies indicate that soil pH may be a critical factor controlling the relative abundances of AOA and AOB communities. In order to determine the relative contributions of AOA and AOB to ammonia oxidation in two agricultural acidic Scottish soils (pH 4.5 and 6), the influence of acetylene (a nitrification inhibitor) was investigated during incubation of soil microcosms at 20 °C for 1 month. High rates of nitrification were observed in both soils in the absence of acetylene. Quantification of respective amoA genes (a key functional gene for ammonia oxidizers) demonstrated significant growth of AOA, but not AOB. A significant positive relationship was found between nitrification rate and AOA, but not AOB growth. AOA growth was inhibited in the acetylene-containing microcosms. Moreover, AOA transcriptional activity decreased significantly in the acetylene-containing microcosms compared with the control, whereas no difference was observed for the AOB transcriptional activity. Consequently, growth and activity of only archaeal but not bacterial ammonia oxidizer communities strongly suggest that AOA, but not AOB, control nitrification in these two acidic soils.

1995 - The New England journal of medicine
A preliminary study of ritonavir, an inhibitor of HIV-1 protease, to treat HIV-1 infection.

BACKGROUND Ritonavir is a potent inhibitor in vitro of human immunodeficiency virus type 1 (HIV-1) protease, which is needed for virions to mature and become infective. We assessed the safety and efficacy of ritonavir in patients with HIV-1 infection. METHODS We administered ritonavir orally to 62 patients in one of four dosages during a 12-week trial containing a 4-week randomized, placebo-controlled, double-blinded phase followed by an 8-week dose-blinded phase. We assessed the response with serial measurements of plasma viremia and serial CD4 cell counts. RESULTS Fifty-two patients completed the 12-week trial. Diarrhea and nausea were the most common side effects, and reversible elevations in serum triglyceride and gamma-glutamyltransferase levels were the most frequent laboratory abnormalities. Ritonavir had a rapid antiviral effect, with a mean maximal reduction in the number of copies of HIV-1 RNA per milliliter of plasma that ranged from 0.86 to 1.18 log in the four dosage groups. After 12 weeks of treatment, the antiviral effect was partially maintained, with a mean reduction in plasma viremia of 0.5 log. When we used a more sensitive assay for HIV-1 RNA in a subgroup of 20 patients, we found that plasma viremia decreased by a mean of 1.7 log. This antiviral effect was partly sustained at week 12, with a mean reduction of approximately 1.1 log. The patients' CD4 cell counts rose during treatment with ritonavir (median increase, 74 and 83 cells per cubic millimeter at weeks 4 and 12, respectively). CONCLUSIONS The protease inhibitor ritonavir is well tolerated and has a potent antiviral effect, as shown by substantial decreases in plasma viremia and significant elevations in CD4 cell counts. Expanded clinical trials of ritonavir are warranted.

1986 - Proceedings of the National Academy of Sciences of the United States of America
Hydrogen peroxide-inducible proteins in Salmonella typhimurium overlap with heat shock and other stress proteins.

Hydrogen peroxide treatment induces the synthesis of 30 proteins in Salmonella typhimurium. Five of these proteins are also induced by heat shock, including the highly conserved DnaK protein. The induction of one of these five proteins by heat shock is dependent on oxyR, a positive regulator of hydrogen peroxide-inducible genes, while the induction of the other four by heat shock is oxyR independent. Five of the 30 hydrogen peroxide-inducible proteins have been identified, and their structural genes have been mapped. Other stresses such as nalidixic acid, ethanol, or cumene hydroperoxide treatment also induce subsets of the 30 hydrogen peroxide-inducible proteins as well as additional proteins. Hydrogen peroxide-inducible proteins are shown to be largely different from those proteins induced by aerobiosis. In addition, the expression of the katG (catalase) gene is shown to be regulated by oxyR at the level of mRNA.

2008 - Cellular and Molecular Life Sciences
The post-transcriptional steps of eukaryotic ribosome biogenesis

Abstract.One of the most important tasks of any cell is to synthesize ribosomes. In eukaryotes, this process occurs sequentially in the nucleolus, the nucleoplasm and the cytoplasm. It involves the transcription and processing of pre-ribosomal RNAs, their proper folding and assembly with ribosomal proteins and the transport of the resulting pre-ribosomal particles to the cytoplasm where final maturation events occur. In addition to the protein and RNA constituents of the mature cytoplasmic ribosomes, this intricate process requires the intervention of numerous protein and small RNA trans-acting factors. These transiently interact with pre-ribosomal particles at various stages of their maturation. Most of the constituents of pre-ribosomal particles have probably now been identified and research in the field is starting to unravel the timing of their intervention and their precise mode of action. Moreover, quality control mechanisms are being discovered that monitor ribosome synthesis and degrade the RNA components of defective pre-ribosomal particles.

2012 - Science
miRNA-Mediated Gene Silencing by Translational Repression Followed by mRNA Deadenylation and Decay

Translation Block MicroRNAs (miRNAs) are small, noncoding RNA genes that are found in the genomes of most eukaryotes, where they play an important role in the regulation of gene expression. Although whether gene activity is repressed by blocking translation of messenger RNA (mRNA) targets, or by promoting their deadenylation and then degradation, has been open to debate. Bazzini et al. (p. 233, published online 15 March) and Djuranovic et al. (p. 237) looked at early points in the repression reaction in the zebrafish embryo or in Drosophila tissue culture cells, respectively, and found that translation was blocked before target mRNAs were significantly deadenylated and degraded. Thus, miRNAs appear to interfere with the initiation step of translation. MicroRNAs act to destroy their messenger RNA targets by blocking translation initiation, which leads to degradation. microRNAs (miRNAs) regulate gene expression through translational repression and/or messenger RNA (mRNA) deadenylation and decay. Because translation, deadenylation, and decay are closely linked processes, it is important to establish their ordering and thus to define the molecular mechanism of silencing. We have investigated the kinetics of these events in miRNA-mediated gene silencing by using a Drosophila S2 cell-based controllable expression system and show that mRNAs with both natural and engineered 3′ untranslated regions with miRNA target sites are first subject to translational inhibition, followed by effects on deadenylation and decay. We next used a natural translational elongation stall to show that miRNA-mediated silencing inhibits translation at an early step, potentially translation initiation.

2001 - Endocrine-related cancer
The epidermal growth factor receptor family as a central element for cellular signal transduction and diversification.

Homeostasis of multicellular organisms is critically dependent on the correct interpretation of the plethora of signals which cells are exposed to during their lifespan. Various soluble factors regulate the activation state of cellular receptors which are coupled to a complex signal transduction network that ultimately generates signals defining the required biological response. The epidermal growth factor receptor (EGFR) family of receptor tyrosine kinases represents both key regulators of normal cellular development as well as critical players in a variety of pathophysiological phenomena. The aim of this review is to give a broad overview of signal transduction networks that are controlled by the EGFR superfamily of receptors in health and disease and its application for target-selective therapeutic intervention. Since the EGFR and HER2 were recently identified as critical players in the transduction of signals by a variety of cell surface receptors, such as G-protein-coupled receptors and integrins, our special focus is the mechanisms and significance of the interconnectivity between heterologous signalling systems.

2000 - Science's STKE
The Interleukin-1 Receptor/Toll-Like Receptor Superfamily: Signal Transduction During Inflammation and Host Defense

The signal transduction pathways activated by the proinflammatory cytokine interleukin-1 (IL-1) have been the focus of much attention because of the important role that IL-1 plays in inflammatory diseases. A number of proteins have been described that participate in the post-receptor activation of the transcription factor nuclear factor κB (NF-κB), and stress-activated protein kinases such as p38 mitogen-activated protein kinase (MAPK). It has also emerged that the type I IL-1 receptor (IL-1RI) is a member of an expanding receptor superfamily. These related receptors all have sequence similarity in their cytosolic regions. The family includes the Drosophila melanogaster protein Toll, the IL-18 receptor (IL-18R), and 10 Toll-like receptors (TLRs), TLR-1 to TLR-10, which bind to microbial products, activating host defense responses. Because of the similarity of IL-1RI to Toll, the conserved sequence in the cytosolic region of these proteins has been termed the Toll-IL-1 receptor (TIR) domain. The same proteins activated during signaling by IL-1RI also participate in signaling by other receptors with TIR domains. The receptor superfamily is evolutionarily conserved; members also occur in plants and insects, where they also function in host defense. The signaling proteins that are activated are also conserved across species. Differences are, however, starting to emerge in signaling pathways activated by different receptors. This receptor superfamily, therefore, represents an ancient signaling system that is a critical determinant of the innate immune and inflammatory responses. The interleukin-1 receptor/Toll-like receptor (IL-1R/TLR) superfamily plays a central role in inflammation and the host response to bacterial infection. It includes receptors for the cytokines IL-1 and IL-18, which are secreted during inflammation and during infection in response to microbial products. Products released by a diverse range of microbes act through TLRs, a notable example being lipopolysaccharide (LPS) from gram-negative bacteria, which stimulates TLR-4. Recognition of such microbes by TLRs provokes host defenses against these microbes. Members of the TLR family occur in species from diverse taxa, including mammals, plants, and insects. In insects, receptors such as Toll are also involved in development. The Toll-IL-1 receptor (TIR) domain occurs in the cytosolic region of all superfamily members and triggers the activation of transcription factors, such as nuclear factor κB (NF-κB). It also activates the mitogen-activated protein kinases (MAPKs) p38 and c-Jun NH2-terminal kinase (JNK). These signals lead to increased expression of proteins involved in immunity and inflammation, resulting in defenses against infectious agents or to chronic inflammation in autoimmune diseases. NF-κB binding sites occur in the promoters of a large number of immune genes, and the signaling pathway to NF-κB is known in detail. Also being uncovered are differences in signaling between TLRs that lead to the tailoring of innate immune responses to the provoking pathogen. The IL-1R/TLR system is therefore evolutionarily conserved and is a critical determinant of innate immune and inflammatory responses.

1993 - Journal of general microbiology
The phylogeny of autotrophic ammonia-oxidizing bacteria as determined by analysis of 16S ribosomal RNA gene sequences.

Partial sequences of the 16S ribosomal RNA genes of eleven autotrophic ammonia-oxidizing bacteria were determined by PCR amplification from small amounts of heat-lysed biomass followed by direct sequencing of PCR products. The sequences were aligned with those of representative Proteobacteria and phylogenetic trees inferred using both parsimony and distance matrix methods. This confirmed that the autotrophic ammonia-oxidizers comprise two major lines of descent within the Proteobacteria. Nitrosomonas spp., Nitrosococcus mobilis, and strains of Nitrosovibrio, Nitrosospira and Nitrosolobus were located in the beta-subdivision. The recovery of Nitrosococcus oceanus strains as a deep branch in the gamma-subdivision supported the RNA catalogue data which had indicated that the genus Nitrosococcus is polyphyletic. The autotrophic ammonia-oxidizing bacteria of the beta-Proteobacteria formed a coherent group which is interpreted as representing a single family. Within this clade, the genera Nitrosovibrio, Nitrosospira and Nitrosolobus exhibited very high levels of homology in their 16S ribosomal RNA gene sequences and can be accommodated within a single genus. Separation of these genera is currently based entirely on gross morphological differences and these can now be considered more appropriate for the identification of species within this group. It is therefore proposed that Nitrosolobus, Nitrosovibrio and Nitrosospira strains be reclassified in a single genus for which the name Nitrosospira has priority.

2006 - Molecular plant-microbe interactions : MPMI
Organic acids, sugars, and L-tryptophane in exudates of vegetables growing on stonewool and their effects on activities of rhizosphere bacteria.

The influence of stonewool substrate on the exudation of the major soluble carbon nutrients and of the auxin precursor tryptophane for Pseudomonas biocontrol agents was studied. To this end, the composition of the organic acids and sugars, as well that of tryptophane, of axenically collected exudates of seed, seedlings, and roots of tomato, cucumber, and sweet pepper was determined. The major results were as follows. i) The total amount of organic acid is much higher than that of total sugar. ii) Exudation of both organic acids and sugars increases during plant growth. iii) Citric, succinic, and malic acids represent the major organic acids, whereas fructose and glucose are the major sugars. iv) Compared with glass beads as a neutral substrate, stonewool substantially stimulates exudation of organic acids and sugars. v) It appeared that enhanced root-tip-colonizing bacteria isolated previously from the rhizosphere of tomato and cucumber grow much better in minimal medium with citrate as the sole carbon source than other, randomly selected rhizobacteria do. This indicates that the procedure which selects for excellent root-tip colonizers enriches for strains which utilize the major exudate carbon source citrate. vi) The content of L-tryptophane, the direct precursor of auxin, is approximately 60-fold higher in seedling exudates of tomato and sweet pepper than in cucumber seedling exudates, indicating a higher possibility of plant growth stimulation after inoculation with auxin-producing rhizobacteria for tomato and sweet pepper crops than for cucumber. However, the biocontrol strain Pseudomonas fluorescens WCS365, which is able to convert tryptophane into auxin, did not stimulate growth of these three crops. In contrast, this strain did stimulate growth of roots of radish, a plant which exudes nine times more tryptophane than tomato does.

1985 - Nature
Control of eukaryotic messenger RNA synthesis by sequence-specific DNA-binding proteins

The enzymatic machinery that carries out RNA synthesis provides the cell with the means to adjust the patterns of transcription in response to environmental and developmental signals. In eukaryotes, this regulation is mediated in part by promoter-specific transcription factors, which are DNA-binding proteins with the ability to discriminate between distinctive DNA sequence elements found in the promoter regions of different genes. The presence of these factors bound to DNA enables other components of the transcriptional machinery, including the RNA polymerase, to initiate transcription with selectivity and accuracy.

1999 - Circulation
Relationship between TIMI frame count and clinical outcomes after thrombolytic administration. Thrombolysis In Myocardial Infarction (TIMI) Study Group.

BACKGROUND The corrected TIMI frame count (CTFC) is the number of cine frames required for dye to first reach standardized distal coronary landmarks, and it is an objective and quantitative index of coronary blood flow. METHODS AND RESULTS The CTFC was measured in 1248 patients in the TIMI 4, 10A, and 10B trials, and its relationship to clinical outcomes was examined. Patients who died in the hospital had a higher CTFC (ie, slower flow) than survivors (69. 6+/-35.4 [n=53] versus 49.5+/-32.3 [n=1195]; P=0.0003). Likewise, patients who died by 30 to 42 days had higher CTFCs than survivors (66.2+/-36.4 [n=57] versus 49.9+/-32.1 [n=1059]; P=0.006). In a multivariate model that excluded TIMI flow grades, the 90-minute CTFC was an independent predictor of in-hospital mortality (OR=1.21 per 10-frame rise [95% CI, 1.1 to 1.3], an approximately 0.7% increase in absolute mortality for every 10-frame rise; P<0.001) even when other significant correlates of mortality (age, heart rate, anterior myocardial infarction, and female sex) were adjusted for in the model. The CTFC identified a subgroup of patients with TIMI grade 3 flow who were at a particularly low risk of adverse outcomes. The risk of in-hospital mortality increased in a stepwise fashion from 0.0% (n=41) in patients with a 90-minute CTFC that was faster than the 95% CI for normal flow (0 to 13 frames, hyperemia, TIMI grade 4 flow), to 2.7% (n=18 of 658 patients) in patients with a CTFC of 14 to 40 (a CTFC of 40 has previously been identified as the cutpoint for distinguishing TIMI grade 3 flow), to 6.4% (35/549) in patients with a CTFC >40 (P=0.003). Although the risk of death, recurrent myocardial infarction, shock, congestive heart failure, or left ventricular ejection fraction </=40% was 13.0% among patients with TIMI grade 3 flow (CTFC </=40), the CTFC tended to segregate patients into lower-risk (CTFC </=20, risk of adverse outcome of 7. 9%) and higher-risk subgroups (CTFC >20 to </=40, risk of adverse outcome of 15.5%; P=0.17). CONCLUSIONS Faster (lower) 90-minute CTFCs are related to improved in-hospital and 1-month clinical outcomes after thrombolytic administration in both univariate and multivariate models. Even among those patients classified as having normal flow (TIMI grade 3 flow, CTFC </=40), there may be lower- and higher-risk subgroups.

2010 - Nature
Cis Interactions between Notch and Delta Generate Mutually Exclusive Signaling States

The Notch–Delta signalling pathway allows communication between neighbouring cells during development. It has a critical role in the formation of ‘fine-grained’ patterns, generating distinct cell fates among groups of initially equivalent neighbouring cells and sharply delineating neighbouring regions in developing tissues. The Delta ligand has been shown to have two activities: it transactivates Notch in neighbouring cells and cis-inhibits Notch in its own cell. However, it remains unclear how Notch integrates these two activities and how the resulting system facilitates pattern formation. Here we report the development of a quantitative time-lapse microscopy platform for analysing Notch–Delta signalling dynamics in individual mammalian cells, with the aim of addressing these issues. By controlling both cis- and trans-Delta concentrations, and monitoring the dynamics of a Notch reporter, we measured the combined cis–trans input–output relationship in the Notch–Delta system. The data revealed a striking difference between the responses of Notch to trans- and cis-Delta: whereas the response to trans-Delta is graded, the response to cis-Delta is sharp and occurs at a fixed threshold, independent of trans-Delta. We developed a simple mathematical model that shows how these behaviours emerge from the mutual inactivation of Notch and Delta proteins in the same cell. This interaction generates an ultrasensitive switch between mutually exclusive sending (high Delta/low Notch) and receiving (high Notch/low Delta) signalling states. At the multicellular level, this switch can amplify small differences between neighbouring cells even without transcription-mediated feedback. This Notch–Delta signalling switch facilitates the formation of sharp boundaries and lateral-inhibition patterns in models of development, and provides insight into previously unexplained mutant behaviours.

1996 - Science
Structure of the A Site of Escherichia coli 16S Ribosomal RNA Complexed with an Aminoglycoside Antibiotic

Aminoglycoside antibiotics that bind to 30S ribosomal A-site RNA cause misreading of the genetic code and inhibit translocation. The aminoglycoside antibiotic paromomycin binds specifically to an RNA oligonucleotide that contains the 30S subunit A site, and the solution structure of the RNA-paromomycin complex was determined by nuclear magnetic resonance spectroscopy. The antibiotic binds in the major groove of the model A-site RNA within a pocket created by an A-A base pair and a single bulged adenine. Specific interactions occur between aminoglycoside chemical groups important for antibiotic activity and conserved nucleotides in the RNA. The structure explains binding of diverse aminoglycosides to the ribosome, their specific activity against prokaryotic organisms, and various resistance mechanisms, and provides insight into ribosome function.

2011 - Gastroenterology
Bile acid is a host factor that regulates the composition of the cecal microbiota in rats.

BACKGROUND & AIMS Alterations in the gastrointestinal microbiota have been associated with metabolic diseases. However, little is known about host factors that induce changes in gastrointestinal bacterial populations. We investigated the role of bile acids in this process because of their strong antimicrobial activities, specifically the effects of cholic acid administration on the composition of the gut microbiota in a rat model. METHODS Rats were fed diets supplemented with different concentrations of cholic acid for 10 days. We used 16S ribosomal RNA gene clone library sequencing and fluorescence in situ hybridization to characterize the composition of the cecal microbiota of the different diet groups. Bile acids in feces, organic acids in cecal contents, and some blood parameters were also analyzed. RESULTS Administration of cholic acid induced phylum-level alterations in the composition of the gut microbiota; Firmicutes predominated at the expense of Bacteroidetes. Cholic acid feeding simplified the composition of the microbiota, with outgrowth of several bacteria in the classes Clostridia and Erysipelotrichi. Externally administered cholic acid was efficiently transformed into deoxycholic acid by a bacterial 7α-dehydroxylation reaction. Serum levels of adiponectin decreased significantly in rats given the cholic acid diet. CONCLUSIONS Cholic acid regulates the composition of gut microbiota in rats, inducing similar changes to those induced by high-fat diets. These findings improve our understanding of the relationship between metabolic diseases and the composition of the gastrointestinal microbiota.

1995 - Microbiological reviews
Nitrogen control in bacteria.

Nitrogen metabolism in prokaryotes involves the coordinated expression of a large number of enzymes concerned with both utilization of extracellular nitrogen sources and intracellular biosynthesis of nitrogen-containing compounds. The control of this expression is determined by the availability of fixed nitrogen to the cell and is effected by complex regulatory networks involving regulation at both the transcriptional and posttranslational levels. While the most detailed studies to date have been carried out with enteric bacteria, there is a considerable body of evidence to show that the nitrogen regulation (ntr) systems described in the enterics extend to many other genera. Furthermore, as the range of bacteria in which the phenomenon of nitrogen control is examined is being extended, new regulatory mechanisms are also being discovered. In this review, we have attempted to summarize recent research in prokaryotic nitrogen control; to show the ubiquity of the ntr system, at least in gram-negative organisms; and to identify those areas and groups of organisms about which there is much still to learn.

2004 - Cell
Regulation of RNA Polymerase II Transcription by Sequence-Specific DNA Binding Factors

In eukaryotes, transcription of the diverse array of tens of thousands of protein-coding genes is carried out by RNA polymerase II. The control of this process is predominantly mediated by a network of thousands of sequence-specific DNA binding transcription factors that interpret the genetic regulatory information, such as in transcriptional enhancers and promoters, and transmit the appropriate response to the RNA polymerase II transcriptional machinery. This review will describe some early advances in the discovery and characterization of the sequence-specific DNA binding transcription factors as well as some of the properties of these regulatory proteins.

1985 - Proceedings of the National Academy of Sciences of the United States of America
A bacteriophage T7 RNA polymerase/promoter system for controlled exclusive expression of specific genes.

The RNA polymerase gene of bacteriophage T7 has been cloned into the plasmid pBR322 under the inducible control of the lambda PL promoter. After induction, T7 RNA polymerase constitutes 20% of the soluble protein of Escherichia coli, a 200-fold increase over levels found in T7-infected cells. The overproduced enzyme has been purified to homogeneity. During extraction the enzyme is sensitive to a specific proteolysis, a reaction that can be prevented by a modification of lysis conditions. The specificity of T7 RNA polymerase for its own promoters, combined with the ability to inhibit selectively the host RNA polymerase with rifampicin, permits the exclusive expression of genes under the control of a T7 RNA polymerase promoter. We describe such a coupled system and its use to express high levels of phage T7 gene 5 protein, a subunit of T7 DNA polymerase.

Signal Transduction and Regulatory Mechanisms Involved in Control of the σS (RpoS) Subunit of RNA Polymerase

R. Hengge-aronis
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R. Hengge‐Aronis

Abstract:SUMMARY The σS (RpoS) subunit of RNA polymerase is the master regulator of the general stress response in Escherichia coli and related bacteria. While rapidly growing cells contain very little σS, exposure to many different stress conditions results in rapid and strong σS induction. Consequently, transcription of numerous σS-dependent genes is activated, many of which encode gene products with stress-protective functions. Multiple signal integration in the control of the cellular σS level is achieved by rpoS transcriptional and translational control as well as by regulated σS proteolysis, with various stress conditions differentially affecting these levels of σS control. Thus, a reduced growth rate results in increased rpoS transcription whereas high osmolarity, low temperature, acidic pH, and some late-log-phase signals stimulate the translation of already present rpoS mRNA. In addition, carbon starvation, high osmolarity, acidic pH, and high temperature result in stabilization of σS, which, under nonstress conditions, is degraded with a half-life of one to several minutes. Important cis-regulatory determinants as well as trans-acting regulatory factors involved at all levels of σS regulation have been identified. rpoS translation is controlled by several proteins (Hfq and HU) and small regulatory RNAs that probably affect the secondary structure of rpoS mRNA. For σS proteolysis, the response regulator RssB is essential. RssB is a specific direct σS recognition factor, whose affinity for σS is modulated by phosphorylation of its receiver domain. RssB delivers σS to the ClpXP protease, where σS is unfolded and completely degraded. This review summarizes our current knowledge about the molecular functions and interactions of these components and tries to establish a framework for further research on the mode of multiple signal input into this complex regulatory system.

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