Genomes contain both a genetic code specifying amino acids and a regulatory code specifying transcription factor (TF) recognition sequences. We used genomic deoxyribonuclease I footprinting to map nucleotide resolution TF occupancy across the human exome in 81 diverse cell types. We found that ~15% of human codons are dual-use codons ("duons") that simultaneously specify both amino acids and TF recognition sites. Duons are highly conserved and have shaped protein evolution, and TF-imposed constraint appears to be a major driver of codon usage bias. Conversely, the regulatory code has been selectively depleted of TFs that recognize stop codons. More than 17% of single-nucleotide variants within duons directly alter TF binding. Pervasive dual encoding of amino acid and regulatory information appears to be a fundamental feature of genome evolution.

BACKGROUND

The specific role of apoptosis in human atherosclerosis remains unknown. During apoptotic cell death, phosphatidylserine exposure on the cell surface confers a high tissue-factor (TF)-dependent procoagulant activity.

RESULTS

In this study, we examined the role of apoptotic cell death in the promotion of plaque thrombogenicity. TF expression and its relation to apoptosis was analyzed in 16 human atherosclerotic plaques by the use of immunohistochemical techniques. The presence of shed membrane apoptotic microparticles was analyzed in extracts from 6 human atherosclerotic plaques and 3 underlying arterial walls. The microparticles were captured by annexin V and their amounts estimated with respect to their phospholipid content by use of a prothrombinase assay. The prothrombogenic potential of the microparticles was further assessed by the measurement of total and microparticle-dependent TF activity in the extracts. The cell origin of the microparticles was determined after capture by specific antibodies. We were able to detect marked TF expression in the plaques in close proximity to apoptotic cells and debris, suggesting a potential interaction between TF and the apoptotic cell surfaces. High levels of shed membrane apoptotic microparticles were detected in extracts from atherosclerotic plaques but not in the underlying arterial walls (29.5+/-3.7 nmol/L phosphatidylserine equivalent versus 1.3+/-0.4 nmol/L, respectively, P<0.02). The microparticles were mainly of monocytic and lymphocytic origin and retained 97+/-2% of total TF activity, indicating a direct causal relationship between shed membrane microparticles and procoagulant activity of plaque extracts.

CONCLUSIONS

These results indicate that shed membrane microparticles with procoagulant potential are produced in human atherosclerotic plaques. Apoptosis could be a critical determinant of plaque thrombogenicity after plaque rupture.

In order to determine how signaling pathways differentially regulate gene expression, it is necessary to identify the interactions between transcription factors (TFs) and their cognate cis-regulatory DNA elements. Here, we have outlined a chromatin immunoprecipitation (ChIP) protocol for use in whole Caenorhabditis elegans extracts. We discuss optimization of the procedure, including growth and harvesting of the worms, formaldehyde fixation, TF immunoprecipitation and analysis of bound sequences through real-time PCR. It takes approximately 10-12 d to obtain the worm culture for ChIP; the ChIP procedure is spaced out over a period of 2.5 d with two overnight incubations.

Task-free functional magnetic resonance imaging (TF-fMRI) has great potential for advancing the understanding and treatment of neurologic illness. However, as with all measures of neural activity, variability is a hallmark of intrinsic connectivity networks (ICNs) identified by TF-fMRI. This variability has hampered efforts to define a robust metric of connectivity suitable as a biomarker for neurologic illness. We hypothesized that some of this variability rather than representing noise in the measurement process, is related to a fundamental feature of connectivity within ICNs, which is their non-stationary nature. To test this hypothesis, we used a large (n = 892) population-based sample of older subjects to construct a well characterized atlas of 68 functional regions, which were categorized based on independent component analysis network of origin, anatomical locations, and a functional meta-analysis. These regions were then used to construct dynamic graphical representations of brain connectivity within a sliding time window for each subject. This allowed us to demonstrate the non-stationary nature of the brain's modular organization and assign each region to a "meta-modular" group. Using this grouping, we then compared dwell time in strong sub-network configurations of the default mode network (DMN) between 28 subjects with Alzheimer's dementia and 56 cognitively normal elderly subjects matched 1:2 on age, gender, and education. We found that differences in connectivity we and others have previously observed in Alzheimer's disease can be explained by differences in dwell time in DMN sub-network configurations, rather than steady state connectivity magnitude. DMN dwell time in specific modular configurations may also underlie the TF-fMRI findings that have been described in mild cognitive impairment and cognitively normal subjects who are at risk for Alzheimer's dementia.

Gene expression is controlled by transcription factors (TFs) that consist of DNA-binding domains (DBDs) and activation domains (ADs). The DBDs have been well characterized, but little is known about the mechanisms by which ADs effect gene activation. Here, we report that diverse ADs form phase-separated condensates with the Mediator coactivator. For the OCT4 and GCN4 TFs, we show that the ability to form phase-separated droplets with Mediator in vitro and the ability to activate genes in vivo are dependent on the same amino acid residues. For the estrogen receptor (ER), a ligand-dependent activator, we show that estrogen enhances phase separation with Mediator, again linking phase separation with gene activation. These results suggest that diverse TFs can interact with Mediator through the phase-separating capacity of their ADs and that formation of condensates with Mediator is involved in gene activation.

The YEAst Search for Transcriptional Regulators And Consensus Tracking (YEASTRACT) information system (http://www.yeastract.com) was developed to support the analysis of transcription regulatory associations in Saccharomyces cerevisiae. Last updated in June 2010, this database contains over 48,200 regulatory associations between transcription factors (TFs) and target genes, including 298 specific DNA-binding sites for 110 characterized TFs. All regulatory associations stored in the database were revisited and detailed information on the experimental evidences that sustain those associations was added and classified as direct or indirect evidences. The inclusion of this new data, gathered in response to the requests of YEASTRACT users, allows the user to restrict its queries to subsets of the data based on the existence or not of experimental evidences for the direct action of the TFs in the promoter region of their target genes. Another new feature of this release is the availability of all data through a machine readable web-service interface. Users are no longer restricted to the set of available queries made available through the existing web interface, and can use the web service interface to query, retrieve and exploit the YEASTRACT data using their own implementation of additional functionalities. The YEASTRACT information system is further complemented with several computational tools that facilitate the use of the curated data when answering a number of important biological questions. Since its first release in 2006, YEASTRACT has been extensively used by hundreds of researchers from all over the world. We expect that by making the new data and services available, the system will continue to be instrumental for yeast biologists and systems biology researchers.

To examine transcription factor (TF) network(s), we created mouse ESC lines, in each of which 1 of 50 TFs tagged with a FLAG moiety is inserted into a ubiquitously controllable tetracycline-repressible locus. Of the 50 TFs, Cdx2 provoked the most extensive transcriptome perturbation in ESCs, followed by Esx1, Sox9, Tcf3, Klf4, and Gata3. ChIP-Seq revealed that CDX2 binds to promoters of upregulated target genes. By contrast, genes downregulated by CDX2 did not show CDX2 binding but were enriched with binding sites for POU5F1, SOX2, and NANOG. Genes with binding sites for these core TFs were also downregulated by the induction of at least 15 other TFs, suggesting a common initial step for ESC differentiation mediated by interference with the binding of core TFs to their target genes. These ESC lines provide a fundamental resource to study biological networks in ESCs and mice.

Understanding how the expression of transcription factor (TF) genes is modulated is essential for reconstructing gene regulatory networks. There is increasing evidence that sequences other than upstream noncoding can contribute to modulating gene expression, but how frequently they do so remains unclear. Here, we investigated the regulation of TFs expressed in a tissue-enriched manner in Arabidopsis roots. For 61 TFs, we created GFP reporter constructs driven by each TF's upstream noncoding sequence (including the 5'UTR) fused to the GFP reporter gene alone or together with the TF's coding sequence. We compared the visually detectable GFP patterns with endogenous mRNA expression patterns, as defined by a genome-wide microarray root expression map. An automated image analysis method for quantifying GFP signals in different tissues was developed and used to validate our visual comparison method. From these combined analyses, we found that (i) the upstream noncoding sequence was sufficient to recapitulate the mRNA expression pattern for 80% (35/44) of the TFs, and (ii) 25% of the TFs undergo posttranscriptional regulation via microRNA-mediated mRNA degradation (2/24) or via intercellular protein movement (6/24). The results suggest that, for Arabidopsis TFs, upstream noncoding sequences are major contributors to mRNA expression pattern establishment, but modulation of transcription factor protein expression pattern after transcription is relatively frequent. This study provides a systematic overview of regulation of TF expression at a cellular level.

To determine the locus, full extent, and topographic organization of cortical connections of area V4 (visual area 4), we injected anterograde and retrograde tracers under electrophysiological guidance into 21 sites in 9 macaques. Injection sites included representations ranging from central to far peripheral eccentricities in the upper and lower fields. Our results indicated that all parts of V4 are connected with occipital areas V2 (visual area 2), V3 (visual area 3), and V3A (visual complex V3, part A), superior temporal areas V4t (V4 transition zone), MT (medial temporal area), and FST (fundus of the superior temporal sulcus [STS] area), inferior temporal areas TEO (cytoarchitectonic area TEO in posterior inferior temporal cortex) and TE (cytoarchitectonic area TE in anterior temporal cortex), and the frontal eye field (FEF). By contrast, mainly peripheral field representations of V4 are connected with occipitoparietal areas DP (dorsal prelunate area), VIP (ventral intraparietal area), LIP (lateral intraparietal area), PIP (posterior intraparietal area), parieto-occipital area, and MST (medial STS area), and parahippocampal area TF (cytoarchitectonic area TF on the parahippocampal gyrus). Based on the distribution of labeled cells and terminals, projections from V4 to V2 and V3 are feedback, those to V3A, V4t, MT, DP, VIP, PIP, and FEF are the intermediate type, and those to FST, MST, LIP, TEO, TE, and TF are feedforward. Peripheral field projections from V4 to parietal areas could provide a direct route for rapid activation of circuits serving spatial vision and spatial attention. By contrast, the predominance of central field projections from V4 to inferior temporal areas is consistent with the need for detailed form analysis for object vision.

Enteral nutrition (EN) by means of oral nutritional supplements (ONS) and tube feeding (TF) offers the possibility to increase or to insure nutrient intake in case of insufficient oral food intake. The present guideline is intended to give evidence-based recommendations for the use of ONS and TF in patients with liver disease (LD). It was developed by an interdisciplinary expert group in accordance with officially accepted standards and is based on all relevant publications since 1985. The guideline was discussed and accepted in a consensus conference. EN by means of ONS is recommended for patients with chronic LD in whom undernutrition is very common. ONS improve nutritional status and survival in severely malnourished patients with alcoholic hepatitis. In patients with cirrhosis, TF improves nutritional status and liver function, reduces the rate of complications and prolongs survival. TF commenced early after liver transplantation can reduce complication rate and cost and is preferable to parenteral nutrition. In acute liver failure TF is feasible and used in the majority of patients.

Apolipoprotein CIII (apoCIII), a lipid-binding protein involved in the transport of triglycerides and cholesterol in the plasma, is synthesized primarily in the liver and the intestine. A cis-acting regulatory element, C3P, located at -90 to -66 upstream from the apoCIII gene transcriptional start site (+1), is necessary for maximal expression of the apoCIII gene in human hepatoma (HepG2) and intestinal carcinoma (Caco2) cells. This report shows that three members of the steroid receptor superfamily of transcription factors, hepatocyte nuclear factor 4 (HNF-4), apolipoprotein AI regulatory protein 1 (ARP-1), and Ear3/COUP-TF, act at the C3P site. HNF-4 activates apoCIII gene expression in HepG2 and Caco2 cells, while ARP-1 and Ear3/COUP-TF repress its expression in the same cells. HNF-4 activation is abolished by increasing amounts of ARP-1 or Ear3/COUP-TF, and repression by ARP-1 or Ear3/COUP-TF is alleviated by increasing amounts of HNF-4. HNF-4 and ARP-1 bind with similar affinities to the C3P site, suggesting that their opposing transcriptional effects may be mediated by direct competition for DNA binding. HNF-4 and ARP-1 mRNAs are present within the same cells in the liver and intestine, and protein extracts from hepatic tissue, HepG2, and Caco2 cells contain significantly more HNF-4 than ARP-1 or Ear3/COUP-TF binding activities. These findings suggest that the transcription of the apoCIII gene in vivo is dependent, at least in part, upon the intracellular balance of these positive and negative regulatory factors.

oPOSSUM-3 is a web-accessible software system for identification of over-represented transcription factor binding sites (TFBS) and TFBS families in either DNA sequences of co-expressed genes or sequences generated from high-throughput methods, such as ChIP-Seq. Validation of the system with known sets of co-regulated genes and published ChIP-Seq data demonstrates the capacity for oPOSSUM-3 to identify mediating transcription factors (TF) for co-regulated genes or co-recovered sequences. oPOSSUM-3 is available at http://opossum.cisreg.ca.

Whereas unperturbed endothelial cells provide potent anticoagulant properties, exposure to inflammatory and atherogenic stimuli can rapidly lead to a procoagulant behavior. Because recent studies provide evidence that apoptosis of vascular cells may occur under conditions such as atherosclerosis and inflammation, we investigated whether apoptotic endothelial cells may contribute to the development of a prothrombotic state. In this report, it is shown that both adherent and detached apoptotic human umbilical vein endothelial cells (HUVECs) become procoagulant. Apoptosis was induced by staurosporine, a nonspecific protein kinase inhibitor, or by culture in suspension with serum deprivation. Both methods resulted in similar findings. As assessed by flow cytometric determination of annexin V binding, HUVECs undergoing cell death exhibited typically a more rapid exposure of membrane phosphatidylserine (PS) than DNA fragmentation. Depending on the stage of apoptosis, this redistribution of phospholipids was found to induce an increase of the activity of the intrinsic tenase complex by 25% to 60%. Although apoptotic cells did not show antigenic or functional tissue factor (TF) activity, when preactivated with lipopolysaccharide, TF procoagulant activity increased by 50% to 70%. At 8 hours after apoptosis induction, antigenic thrombomodulin, heparan sulfates, and TF pathway inhibitor decreased by about 83%, 80%, and 59%, respectively. The functional activity of these components was reduced by about 36%, 52%, and 39%, respectively. Moreover, the presence of apoptotic HUVECs led to a significant increase of thrombin formation in recalcified citrated plasma. In conclusion, apoptotic HUVECs, either adherent or in suspension, become procoagulant by increased expression of PS and the loss of anticoagulant membrane components.

Transcription factors (TFs) bind specifically to discrete regions of mammalian genomes called cis-regulatory elements. Among those are enhancers, which play key roles in regulation of gene expression during development and differentiation. Despite the recognized central regulatory role exerted by chromatin in control of TF functions, much remains to be learned regarding the chromatin structure of enhancers and how it is established. Here, we have analyzed on a genomic-scale enhancers that recruit FOXA1, a pioneer transcription factor that triggers transcriptional competency of these cis-regulatory sites. Importantly, we found that FOXA1 binds to genomic regions showing local DNA hypomethylation and that its cell-type-specific recruitment to chromatin is linked to differential DNA methylation levels of its binding sites. Using neural differentiation as a model, we showed that induction of FOXA1 expression and its subsequent recruitment to enhancers is associated with DNA demethylation. Concomitantly, histone H3 lysine 4 methylation is induced at these enhancers. These epigenetic changes may both stabilize FOXA1 binding and allow for subsequent recruitment of transcriptional regulatory effectors. Interestingly, when cloned into reporter constructs, FOXA1-dependent enhancers were able to recapitulate their cell type specificity. However, their activities were inhibited by DNA methylation. Hence, these enhancers are intrinsic cell-type-specific regulatory regions of which activities have to be potentiated by FOXA1 through induction of an epigenetic switch that includes notably DNA demethylation.

Cell cycle progression is a process that is tightly controlled by internal and external signals. Environmental cues, such as those provided by growth factors, activate early signals that promote cell cycle entry. Cells that have progressed past the restriction point become independent of growth factors, and cell cycle progression is then controlled endogenously. The phosphatidylinositol 3OH kinase (PI(3)K)/protein kinase B (PKB) pathway must be activated in G1 to inactivate forkhead transcription factors (FKH-TFs) and allow cell cycle entry. Here we show that subsequent attenuation of the PI(3)K/PKB pathway is required to allow transcriptional activation of FKH-TF in G2. FKH-TF activity in G2 controls mammalian cell cycle termination, as interference with FKH transcriptional activation by disrupting PI(3)K/PKB downregulation, or by expressing a transcriptionally inactive FKH mutant, induces cell accumulation in G2/M, defective cytokinesis, and delayed transition from M to G1 of the cell cycle. We demonstrate that FKH-TFs regulate expression of mitotic genes such as cyclin B and polo-like kinase (Plk). Our results support the important role of forkhead in the control of mammalian cell cycle completion, and suggest that efficient execution of the mitotic programme depends on downregulation of PI(3)K/PKB and consequent induction of FKH transcriptional activity.

The combination of ChIP-seq and transcriptome analysis is a compelling approach to unravel the regulation of gene expression. Several recently published methods combine transcription factor (TF) binding and gene expression for target prediction, but few of them provide an efficient software package for the community. Binding and expression target analysis (BETA) is a software package that integrates ChIP-seq of TFs or chromatin regulators with differential gene expression data to infer direct target genes. BETA has three functions: (i) to predict whether the factor has activating or repressive function; (ii) to infer the factor's target genes; and (iii) to identify the motif of the factor and its collaborators, which might modulate the factor's activating or repressive function. Here we describe the implementation and features of BETA to demonstrate its application to several data sets. BETA requires ~1 GB of RAM, and the procedure takes 20 min to complete. BETA is available open source at http://cistrome.org/BETA/.

Conservation of the vertebrate body plan has been attributed to the evolutionary stability of gene-regulatory networks (GRNs). We describe a regulatory circuit made up of Gata2, Fli1, and Scl/Tal1 and their enhancers, Gata2-3, Fli1+12, and Scl+19, that operates during specification of hematopoiesis in the mouse embryo. We show that the Fli1+12 enhancer, like the Gata2-3 and Scl+19 enhancers, targets hematopoietic stem cells (HSCs) and relies on a combination of Ets, Gata, and E-Box motifs. We show that the Gata2-3 enhancer also uses a similar cluster of motifs and that Gata2, Fli1, and Scl are expressed in embryonic day-11.5 dorsal aorta where HSCs originate and in fetal liver where they multiply. The three HSC enhancers in these tissues and in ES cell-derived hemangioblast equivalents are bound by each of these transcription factors (TFs) and form a fully connected triad that constitutes a previously undescribed example of both this network motif in mammalian development and a GRN kernel operating during the specification of a mammalian stem cell.

Transferrin (Tf) receptor-variant Chinese hamster ovary cells have been isolated by selection for resistance to two Tf-toxin conjugates. The hybrid toxins contain Tf covalently linked to ricin A chain or a genetically engineered diphtheria toxin fragment. The Tf-receptor-variant (TRV) cells do not have detectable cell-surface Tf receptor; they do not bind fluorescein-Tf or 125I-Tf. TRV cells are at least 100-fold more resistant to the Tf-diphtheria toxin conjugate than are the parent cells. The TRV cells have retained sensitivity to native diphtheria toxin, indicating that the increased resistance to the conjugate is correlated with the loss of Tf binding. The endocytosis of fluorescein-labeled alpha 2-macroglobulin is normal in TRV cells, demonstrating that the defect does not pleiotropically affect endocytosis. Since these cells lack endogenous Tf receptor activity, they are ideally suited for studies of the functional expression of normal or altered Tf receptors introduced into the cells by cDNA transfection. One advantage of this system is that Tf binding and uptake can be used to monitor the behavior of the transfected receptor. A cDNA clone of the human Tf receptor has been transfected into TRV cells. In the stably expressing transfectants, the behavior of the human receptor is very similar to that of the endogenous Chinese hamster ovary cell Tf receptor. Tf binds to cell surface receptors, and is internalized into the para-Golgi region of the cell. Iron is released from Tf, and the apo-Tf and its receptor are recycled back to the cell surface. Thus, the TRV cells can be used to study the behavior of genetically altered Tf receptors in the absence of interfering effects from endogenous receptors.

Chitin, found in the cell walls of true fungi and the exoskeleton of insects and nematodes, is a well-established elicitor of plant defense responses. In this study, we analyzed the expression patterns of Arabidopsis thaliana transcription factor (TF) and ubiquitin-ligase genes in response to purified chitooctaose at different treatment times (15, 30, 60, 90, and 120 min after treatment), using both quantitative polymerase chain reaction and the Affymetrix Arabidopsis whole-genome array. A total of 118 TF genes and 30 ubiquitin-ligase genes were responsive to the chitin treatment. Among these genes, members from the following four TF families were overrepresented: APETALA2/ethylene-reponsive element binding proteins (27), C2H2 zinc finger proteins (14), MYB domain-containing proteins (11), and WRKY domain transcription factors (14). Transcript variants from a few of these genes were found to respond differentially to chitin, suggesting transcript-specific regulation of these TF genes.

The epithelial to mesenchymal transition (EMT) is an important step for the developmental process. Recent evidences support that EMT allows the tumor cells to acquire invasive properties and to develop metastatic growth characteristics. Some of the transcription factors, which are actively involved in EMT process, have a significant role in the EMT-metastasis linkage. A number of studies have reported that EMT-inducing transcription factors (EMT-TFs), such as Twist, Snail, Slug, and Zeb, are directly or indirectly involved in cancer cell metastasis through a different signaling cascades, including the Akt, signal transducer and activator of transcription 3 (STAT3), mitogen-activated protein kinase (MAPK) and Wnt pathways, with the ultimate consequence of the downregulation of E-cadherin and upregulation of metastatic proteins, such as N-cadherin, vimentin, matrix metalloproteinase (MMP)-2, etc. This review summarizes the update information on the association of EMT-TFs with cancer metastasis and the possible cancer therapeutics via targeting the EMT-TFs.

BACKGROUND

Plaque disruption with thrombosis commonly causes the acute coronary syndromes. Macrophages, abundant at sites of plaque rupture, release proteinases that weaken plaques and express tissue factor (TF), which initiates thrombosis. The signals that induce expression of these macrophage functions, particularly TF, remain obscure. Recent studies have localized the receptor CD40 and its ligand in human atheroma. This study tested the hypothesis that ligation of CD40 can activate key mononuclear phagocyte functions related to clinical manifestations of atheroma.

RESULTS

Stimulation of human monocytes/macrophages through CD40 by either membranes from activated T cells or recombinant CD40L (rCD40L) induced expression of interstitial collagenase, stromelysin, and TF protein and activity. In contrast, the soluble cytokines interleukin-1 or tumor necrosis factor-alpha did not induce or weakly induced TF expression. Neutralization with anti-CD40L antibody markedly inhibited these actions of both T-cell membranes and rCD40L.

CONCLUSIONS

By inducing the expression of matrix-degrading proteinases and of TF procoagulant, CD40 signaling may contribute to the triggering of acute coronary events.

Individual neuronal cell types are defined by the expression of unique batteries of terminal differentiation genes. The elucidation of the cis-regulatory architecture of several distinct, single neuron type-specific gene batteries in Caenorhabditis elegans has revealed a strikingly simple cis-regulatory logic, in which small cis-regulatory motifs are activated in postmitotic neurons by autoregulating transcription factors (TFs). Loss of the TFs results in the loss of the identity of the individual neuron type. I propose to term these TFs "terminal selector genes" and their cognate cis-regulatory target sites "terminal selector motifs." Terminal selector genes assign individual neuronal identities by directly controlling the expression of downstream, terminal differentiation genes and act in specific regulatory network configurations. The simplicity of the cis-regulatory logic on which the terminal selector gene concept is based may contribute to the evolvability of neuronal diversity.

We examined the intracellular transport of sterol in living cells using a naturally fluorescent cholesterol analog, dehydroergosterol (DHE), which has been shown to mimic many of the properties of cholesterol. By using DHE loaded on methyl-beta-cyclodextrin, we followed this cholesterol analog in pulse-chase studies. At steady state, DHE co-localizes extensively with transferrin (Tf), a marker for the endocytic recycling compartment (ERC), and redistributes with Tf in cells with altered ERC morphology. Expression of a dominant-negative mutation of an ERC-associated protein, mRme-1 (G429R), results in the slowing of both DHE and Tf receptor return to the cell surface. [3H]Cholesterol is found in the same fraction as 125I-Tf on sucrose density gradients, and this fraction can be specifically shifted to a higher density based on the presence of horseradish peroxidase-conjugated Tf in the same organelle. Whereas vesicular transport of Tf and efflux of DHE from the ERC are entirely blocked in energy-depleted cells, delivery of DHE to the ERC from the plasma membrane is only slightly affected. Biochemical studies performed using [3H]cholesterol show that the energy dependence of cholesterol transport to and from the ERC is similar to DHE transport. We propose that a large portion of intracellular cholesterol is localized in the ERC, and this pool might be important in maintaining cellular cholesterol homeostasis.

BACKGROUND

Precise analysis of the genetic network, gene function and transcription regulation requires accurate prediction of transcription factor (TF) bindability on DNA. For calculating the matching score between an input sequence and a set of known TF binding sites, we use positional weight matrices (PWMs) and Bucher's calculating method (Bucher, J. Mol. Biol., 212, 563-578, 1990). Since estimating TF binding sites requires cut-off values, we propose a robust cut-off value determining algorithm.

RESULTS

We generalize the concept of local overrepresentation with statistics, and propose a new algorithm for determining the cut-off value using the background rate estimated on non-promoters. The algorithm iteratively determines parameters separating instances into phenomena-dependent and phenomena-independent subsets. Our system includes the method of re-estimating cut-off values of TFs that mis-recognize other TF preferred regions. Our data source comprised 433 non-redundant vertebrate promoters including viral promoters, from Eukaryotic Promoter Database (EPD) R.50. The method is applied to 205 vertebrate TFs that have frequency matrices in TRANSFAC Ver.3. 4 and the cut-off values of all of them can be determined.

BACKGROUND

The cut-off values and TF binding site predicting tool are available at http://www.hgc.ims.u-tokyo.ac. jp/service/tooldoc/TFBIND. We also provide the cut-off value estimating programs.