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At least 865 records · Page 48Linked to original sources
PubMed2003

Evidence for the adaptive evolution of the carbon fixation gene rbcL during diversification in temperature tolerance of a clade of hot spring cyanobacteria.

Determining the molecular basis of enzyme adaptation is central to understanding the evolution of environmental tolerance but is complicated by the fact that not all amino acid differences between ecologically divergent taxa are adaptive. Analysing patterns of nucleotide sequence evolution can potentially guide the investigation of protein adaptation by identifying candidate codon sites on which diversifying selection has been operating. Here, I test whether there is evidence for molecular adaptation of the carbon fixation gene rbcL for a clade of hot spring cyanobacteria in the genus Synechococcus that has diverged in thermotolerance. Amino acid replacements during Synechococcus radiation have resulted in an increase in the number of hydrophobic residues in the RbcLs of more thermotolerant strains. A similar increase in hydrophobicity has been observed for many thermostable proteins. Maximum likelihood models which allow for heterogeneity among codon sites in the ratio of nonsynonymous to synonymous nucleotide substitutions estimated a class of amino acid sites as a target of positive selection. Depending on the model, a single amino acid site that interacts with a flexible element involved in the opening and closing of the active site was estimated with either low or moderate support to be a member of this class. Site-directed mutagenesis approaches are being explored in order to directly test its adaptive significance.

Adaptation, Biological↗
PubMed2002

GeneChip expression analysis of the iron starvation response in Pseudomonas aeruginosa: identification of novel pyoverdine biosynthesis genes.

Upon iron restriction, the opportunistic pathogen Pseudomonas aeruginosa produces various virulence factors, including siderophores, exotoxin, proteases and haemolysin. The ferric uptake regulator (Fur) plays a central role in this response and also controls other regulatory genes, such as pvdS, which encodes an alternative sigma factor. This circuit leads to a hierarchical cascade of direct and indirect iron regulation. We used the GeneChip to analyse the global gene expression profiles in response to iron. In iron-starved cells,the expression of 118 genes was increased at least fivefold compared with that in iron-replete cells, whereas the expression of 87 genes was decreased at least fivefold. The GeneChip data correlated well with results obtained using individual lacZ gene fusions. Strong iron regulation was observed for previously identified genes involved in biosynthesis or uptake of the siderophores pyoverdine and pyochelin, utilization of heterologous siderophores and haem and ferrous iron transport. A low-iron milieu led to increased expression of the genes encoding TonB, alkaline protease,PrpL protease, exotoxin A, as well as fumarase C, Mn-dependent superoxide dismutase SodA, a ferredoxin and ferredoxin reductase and several oxidoreductases and dehydrogenases. Iron-controlled regulatory genes included seven alternative sigma factors and five other transcriptional regulators. Roughly 20% of the iron-regulated genes encoded proteins of unknown function and lacked any conclusive homologies. Under low-iron conditions, expression of 26 genes or operons was reduced in a DeltapvdS mutant compared with wild type, including numerous novel pyoverdine biosynthetic genes. The GeneChip proved to be a very useful tool for rapid gene expression analysis and identification of novel genes controlled by Fur or PvdS.

Bacterial Proteins↗
PubMed2000

Molecular cloning of geranyl diphosphate synthase and compartmentation of monoterpene synthesis in plant cells.

The nature of isoprenoids synthesized in plants is primarily determined by the specificity of prenyltransferases. Several of these enzymes have been characterized at the molecular level. The compartmentation and molecular regulation of geranyl diphosphate (GPP), the carbon skeleton that is the backbone of myriad monoterpene constituents involved in plant defence, allelopathic interactions and pollination, is poorly understood. We describe here the cloning and functional expression of a GPP synthase (GPPS) from Arabidopsis thaliana. Immunohistological analyses of diverse non-secretory and secretory plant tissues reveal that GPPS and its congeners, monoterpene synthase, deoxy-xylulose phosphate synthase and geranylgeranyl diphosphate synthase, are equally compartmentalized and distributed in non-green plastids as well in chloroplasts of photosynthetic cells. This argues that monoterpene synthesis is not solely restricted to specialized secretory structures but can also occur in photosynthetic parenchyma. These data provide new information as to how monoterpene biosynthesis is compartmentalized and induced de novo in response to biotic and abiotic stress in diverse plants.

Alternative Splicing↗
PubMed2003

Innate and acquired immunity intersect in a global view of the acute-phase response.

Recognition of stereotypic chemical patterns by sentinel cells of the innate immune system provokes a transient deviation from homeostasis, the acute-phase response (APR). Although APR effectors have been identified individually, the complexity of the response suggested that emergent properties would be uncovered by a more comprehensive examination. Our global assessment revealed that approximately 7% of genes in the mouse are mobilized in the hepatic APR to endotoxin. Extensive metabolic adjustments include suppression of pathways for cholesterol, fatty acid, and phospholipid synthesis. Increased expression of genes for innate defense was accompanied by coordinate induction of the MHC class I antigen presentation machinery, illustrating an intersection between innate and adaptive immunity.

Acute-Phase Reaction↗
PubMed2004

Sfp1 is a stress- and nutrient-sensitive regulator of ribosomal protein gene expression.

Yeast cells modulate their protein synthesis capacity in response to physiological needs through the transcriptional control of ribosomal protein (RP) genes. Here we demonstrate that the transcription factor Sfp1, previously shown to play a role in the control of cell size, regulates RP gene expression in response to nutrients and stress. Under optimal growth conditions, Sfp1 is localized to the nucleus, bound to the promoters of RP genes, and helps promote RP gene expression. In response to inhibition of target of rapamycin (TOR) signaling, stress, or changes in nutrient availability, Sfp1 is released from RP gene promoters and leaves the nucleus, and RP gene transcription is down-regulated. Additionally, cells lacking Sfp1 fail to appropriately modulate RP gene expression in response to environmental cues. We conclude that Sfp1 integrates information from nutrient- and stress-responsive signaling pathways to help control RP gene expression.

Cyclic AMP-Dependent Protein Kinases↗
PubMed2005

Independence of replisomes in Escherichia coli chromosomal replication.

In Escherichia coli DNA replication is carried out by the coordinated action of the proteins within a replisome. After replication initiation, the two bidirectionally oriented replisomes from a single origin are colocalized into higher-order structures termed replication factories. The factory model postulated that the two replisomes are also functionally coupled. We tested this hypothesis by using DNA combing and whole-genome microarrays. Nascent DNA surrounding oriC in single, combed chromosomes showed instead that one replisome, usually the leftward one, was significantly ahead of the other 70% of the time. We next used microarrays to follow replication throughout the genome by measuring DNA copy number. We found in multiple E. coli strains that the replisomes are independent, with the leftward replisome ahead of the rightward one. The size of the bias was strain-specific, varying from 50 to 130 kb in the array results. When we artificially blocked one replisome, the other continued unabated, again demonstrating independence. We suggest an improved version of the factory model that retains the advantages of threading DNA through colocalized replisomes at about equal rates, but allows the cell flexibility to overcome obstacles encountered during elongation.

Chromosomes↗
PubMed2006

Microbial diversity in the deep sea and the underexplored "rare biosphere".

The evolution of marine microbes over billions of years predicts that the composition of microbial communities should be much greater than the published estimates of a few thousand distinct kinds of microbes per liter of seawater. By adopting a massively parallel tag sequencing strategy, we show that bacterial communities of deep water masses of the North Atlantic and diffuse flow hydrothermal vents are one to two orders of magnitude more complex than previously reported for any microbial environment. A relatively small number of different populations dominate all samples, but thousands of low-abundance populations account for most of the observed phylogenetic diversity. This "rare biosphere" is very ancient and may represent a nearly inexhaustible source of genomic innovation. Members of the rare biosphere are highly divergent from each other and, at different times in earth's history, may have had a profound impact on shaping planetary processes.

Biodiversity↗
PubMed2002

The hypocholesterolemic agent LY295427 up-regulates INSIG-1, identifying the INSIG-1 protein as a mediator of cholesterol homeostasis through SREBP.

Oxysterols regulate cholesterol homeostasis through liver X receptor (LXR; cholesterol-lowering)- and sterol regulatory element-binding protein (SREBP; cholesterol-raising)-mediated signaling pathways. Previously we reported that the hypocholesterolemic agent LY295427 (4alpha-allylcholestan-3alpha-ol) reverses oxysterol-mediated suppression of SREBP processing. We now report that LY295427 increases expression of insulin-induced gene-1 (INSIG-1) and restores SREBP processing in cells treated with oxysterols. In cells overexpressing the INSIG-1 gene, by contrast, SREBP processing is suppressed and oxysterol regulation is disrupted. SREBP processing is not restored by addition of LY295427, but is restored by increasing the levels of SREBP cleavage-activating protein (SCAP). These findings suggest that the INSIG-1 protein alters sterol balance by modulating SREBP processing jointly with SCAP. To test whether the action of oxysterols on SREBP processing is mediated through endogenous INSIG-1 protein, we used RNAi to lower the expression of the INSIG-1 gene, and found that reduced INSIG-1 protein levels caused the loss of SREBP regulation by oxysterols. We conclude that: (i) INSIG-1 gene expression is suppressed by oxysterols; (ii) LY295427 treatment counters the suppressive effects of oxysterols on SREBP processing, resulting in the expression of the INSIG-1 gene; and (iii) INSIG-1 gene expression affects SREBP processing. Taken together, these data suggest that INSIG-1 plays a critical role in regulating cholesterol concentrations in the cell.

Anticholesteremic Agents↗
PubMed2002

Biliverdin reductase: a major physiologic cytoprotectant.

Bilirubin, an abundant pigment that causes jaundice, has long lacked any clear physiologic role. It arises from enzymatic reduction by biliverdin reductase of biliverdin, a product of heme oxygenase activity. Bilirubin is a potent antioxidant that we show can protect cells from a 10,000-fold excess of H2O2. We report that bilirubin is a major physiologic antioxidant cytoprotectant. Thus, cellular depletion of bilirubin by RNA interference markedly augments tissue levels of reactive oxygen species and causes apoptotic cell death. Depletion of glutathione, generally regarded as a physiologic antioxidant cytoprotectant, elicits lesser increases in reactive oxygen species and cell death. The potent physiologic antioxidant actions of bilirubin reflect an amplification cycle whereby bilirubin, acting as an antioxidant, is itself oxidized to biliverdin and then recycled by biliverdin reductase back to bilirubin. This redox cycle may constitute the principal physiologic function of bilirubin.

Antioxidants↗
PubMed2003

Multiple pathways are co-regulated by the protein kinase Snf1 and the transcription factors Adr1 and Cat8.

ADR1 and CAT8 encode carbon source-responsive transcriptional regulators that cooperatively control expression of genes involved in ethanol utilization. These transcription factors are active only after the diauxic transition, when glucose is depleted and energy-generating metabolism has shifted to the aerobic oxidation of non-fermentable carbon sources. The Snf1 protein kinase complex is required for activation of their downstream target genes described previously. Using DNA microarrays, we determined the extent to which these three factors collaborate in regulating the expression of the yeast genome after glucose depletion. The expression of 108 genes is significantly decreased in the absence of ADR1. The importance of ADR1 during the diauxic transition is illustrated by the observation that expression of almost one-half of the 40 most highly glucose-repressed genes is ADR1-dependent. ADR1-dependent genes fall into a variety of functional classes with carbon metabolism containing the largest number of members. Most of the genes in this class are involved in the oxidation of different non-fermentable carbon sources. These microarray data show that ADR1 coordinates the biochemical pathways that generate acetyl-CoA and NADH from non-fermentable substrates. Only a small number of ADR1-dependent genes are also CAT8-dependent. However, nearly one-half of the ADR1-dependent genes are also dependent on the Snf1 protein kinase for derepression. Many more genes are SNF1-dependent than are either ADR1- or CAT8-dependent suggesting that SNF1 plays a broader role in gene expression than either ADR1 or CAT8. The largest class of SNF1-dependent genes encodes regulatory proteins that could extend SNF1 dependence to additional pathways.

Binding Sites↗
PubMed2003

Coupling of the transcriptional regulation of glutathione biosynthesis to the availability of glutathione and methionine via the Met4 and Yap1 transcription factors.

Depletion of the cellular pool of glutathione is detrimental to eukaryotic cells and in Saccharomyces cerevisiae leads to sensitivity to oxidants and xenobiotics and an eventual cell cycle arrest. Here, we show that the Yap1 and Met4 transcription factors regulate the expression of gamma-glutamylcysteine synthetase (GSH1), encoding the rate-limiting enzyme in glutathione biosynthesis to prevent the damaging effects of glutathione depletion. Transcriptional profiling of a gsh1 mutant indicates that glutathione depletion leads to a general activation of Yap1 target genes, but the expression of Met4-regulated genes remains unaltered. Glutathione depletion appears to result in Yap1 activation via oxidation of thioredoxins, which normally act to down-regulate the Yap1-mediated response. The requirement for Met4 in regulating GSH1 expression is lost in the absence of the centromere-binding protein Cbf1. In contrast, the Yap1-mediated effect is unaffected, indicating that Met4 acts via Cbf1 to regulate the Yap1-mediated induction of GSH1 expression in response to glutathione depletion. Furthermore, yeast cells exposed to the xenobiotic 1-chloro-2,4-dintrobenzene are rapidly depleted of glutathione, accumulate oxidized thioredoxins, and elicit the Yap1/Met4-dependent transcriptional response of GSH1. The addition of methionine, which promotes Met4 ubiquitination and inactivation, specifically represses GSH1 expression after 1-chloro-2,4-dintrobenzene exposure but does not affect Yap1 activation. These results indicate that the Yap1-dependent activation of GSH1 expression in response to glutathione depletion is regulated by the sulfur status of the cell through a specific Met4-dependent mechanism.

Basic Helix-Loop-Helix Leucine Zipper Transcriptio↗
PubMed2004

Connective tissue growth factor (CTGF) is regulated by Wnt and bone morphogenetic proteins signaling in osteoblast differentiation of mesenchymal stem cells.

Osteoblast lineage-specific differentiation of mesenchymal stem cells is a well regulated but poorly understood process. Both bone morphogenetic proteins (BMPs) and Wnt signaling are implicated in regulating osteoblast differentiation and bone formation. Here we analyzed the expression profiles of mesenchymal stem cells stimulated with Wnt3A and osteogenic BMPs, and we identified connective tissue growth factor (CTGF) as a potential target of Wnt and BMP signaling. We confirmed the microarray results, and we demonstrated that CTGF was up-regulated at the early stage of BMP-9 and Wnt3A stimulations and that Wnt3A-regulated CTGF expression was beta-catenin-dependent. RNA interference-mediated knockdown of CTGF expression significantly diminished BMP-9-induced, but not Wnt3A-induced, osteogenic differentiation, suggesting that Wnt3A may also regulate osteoblast differentiation in a CTGF-independent fashion. However, constitutive expression of CTGF was shown to inhibit both BMP-9- and Wnt3A-induced osteogenic differentiation. Exogenous expression of CTGF was shown to promote cell migration and recruitment of mesenchymal stem cells. Our findings demonstrate that CTGF is up-regulated by Wnt3A and BMP-9 at the early stage of osteogenic differentiation, which may regulate the proliferation and recruitment of osteoprogenitor cells; however, CTGF is down-regulated as the differentiation potential of committed pre-osteoblasts increases, strongly suggesting that tight regulation of CTGF expression may be essential for normal osteoblast differentiation of mesenchymal stem cells.

Adenoviridae↗
PubMed2004

p53 suppresses c-Myb-induced trans-activation and transformation by recruiting the corepressor mSin3A.

p53 is known to repress transcription of a number of genes, but the mechanism of p53 recruitment to these target genes is unknown. The c-myb proto-oncogene product (c-Myb) positively regulates proliferation of immature hematopoietic cells, whereas p53 blocks cell cycle progression. Here, we demonstrate that p53 inhibits c-Myb-induced transcription and transformation by directly binding to c-Myb. The ability of c-Myb to maintain the undifferentiated state of M1 cells was also suppressed by p53. p53 did not affect the ability of c-Myb to bind to DNA but formed a ternary complex with the corepressor mSin3A and c-Myb. Thus, p53 antagonizes c-Myb by recruiting mSin3A to down-regulate specific Myb target genes.

Apoptosis↗
PubMed2006

Disruption of an SP2/KLF6 repression complex by SHP is required for farnesoid X receptor-induced endothelial cell migration.

The farnesoid X receptor (FXR) signaling pathway regulates bile acid and cholesterol homeostasis. Here, we demonstrate, using a variety of gain- and loss-of-function approaches, a role of FXR in the process of cell motility, which involves the small heterodimeric partner (SHP)-dependent up-regulation of matrix metalloproteinase-9. We use this observation to reveal a transcriptional regulatory mechanism involving the SP/KLF transcription factors, SP2 and KLF6. Small interference RNA-based silencing studies in combination with promoter, gel shift, and chromatin immunoprecipitation assays indicate that SP2 and KLF6 bind to the matrix metalloproteinase-9 promoter and together function to maintain this gene in a silenced state. However, upon activation of FXR, SHP interacts with SP2 and KLF6, disrupting the SP2/KLF6 repressor complex. Thus, together, these studies identify a mechanism for antagonizing Sp/KLF protein repression function via SHP, with this process regulating endothelial cell motility.

Cell Movement↗
PubMed2002

Complementary profiling of gene expression at the transcriptome and proteome levels in Saccharomyces cerevisiae.

Using an integrated genomic and proteomic approach, we have investigated the effects of carbon source perturbation on steady-state gene expression in the yeast Saccharomyces cerevisiae growing on either galactose or ethanol. For many genes, significant differences between the abundance ratio of the messenger RNA transcript and the corresponding protein product were observed. Insights into the perturbative effects on genes involved in respiration, energy generation, and protein synthesis were obtained that would not have been apparent from measurements made at either the messenger RNA or protein level alone, illustrating the power of integrating different types of data obtained from the same sample for the comprehensive characterization of biological systems and processes.

Databases as Topic↗
PubMed2005

Integrating global proteomic and genomic expression profiles generated from islet alpha cells: opportunities and challenges to deriving reliable biological inferences.

Systematic profiling of expressed gene products represents a promising research strategy for elucidating the molecular phenotypes of islet cells. To this end, we have combined complementary genomic and proteomic methods to better assess the molecular composition of murine pancreatic islet glucagon-producing alphaTC-1 cells as a model system, with the expectation of bypassing limitations inherent to either technology alone. Gene expression was measured with an Affymetrix MG_U74Av2 oligonucleotide array, while protein expression was examined by performing high-resolution gel-free shotgun MS/MS on a nuclear-enriched cell extract. Both analyses were carried out in triplicate to control for experimental variability. Using a stringent detection p value cutoff of 0.04, 48% of all potential mRNA transcripts were predicted to be expressed (probes classified as present in at least two of three replicates), while 1,651 proteins were identified with high-confidence using rigorous database searching. Although 762 of 888 cross-referenced cognate mRNA-protein pairs were jointly detected by both platforms, a sizeable number (126) of gene products was detected exclusively by MS alone. Conversely, marginal protein identifications often had convincing microarray support. Based on these findings, we present an operational framework for both interpreting and integrating dual genomic and proteomic datasets so as to obtain a more reliable perspective into islet alpha cell function.

Animals↗
PubMed2002

Historical development of the concept of the gene.

The classical view of the gene prevailing during the 1910s and 1930s comprehended the gene as the indivisible unit of genetic transmission, genetic recombination, gene mutation and gene function. The discovery of intragenic recombination in the early 1940s led to the neoclassical concept of the gene, which prevailed until the 1970s. In this view the gene or cistron, as it was now called, was divided into its constituent parts, the mutons and recons, materially identified as nucleotides. Each cistron was believed to be responsible for the synthesis of one single mRNA and concurrently for one single polypeptide. The discoveries of DNA technology, beginning in the early 1970s, have led to the second revolution in the concept of the gene in which none of the classical or neoclassical criteria for the definition of the gene hold strictly true. These are the discoveries concerning gene repetition and overlapping, movable genes, complex promoters, multiple polyadenylation sites, polyprotein genes, editing of the primary transcript, pseudogenes and gene nesting. Thus, despite the fact that our comprehension of the structure and organization of the genetic material has greatly increased, we are left with a rather abstract, open and general concept of the gene. This article discusses past and present contemplations of genes, genomes, genotypes and phenotypes as well as the most recent advances of the study of the organization of genomes.

Genetics↗