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1.  The Next Generation of Transcription Factor Binding Site Prediction 
PLoS Computational Biology  2013;9(9):e1003214.
Finding where transcription factors (TFs) bind to the DNA is of key importance to decipher gene regulation at a transcriptional level. Classically, computational prediction of TF binding sites (TFBSs) is based on basic position weight matrices (PWMs) which quantitatively score binding motifs based on the observed nucleotide patterns in a set of TFBSs for the corresponding TF. Such models make the strong assumption that each nucleotide participates independently in the corresponding DNA-protein interaction and do not account for flexible length motifs. We introduce transcription factor flexible models (TFFMs) to represent TF binding properties. Based on hidden Markov models, TFFMs are flexible, and can model both position interdependence within TFBSs and variable length motifs within a single dedicated framework. The availability of thousands of experimentally validated DNA-TF interaction sequences from ChIP-seq allows for the generation of models that perform as well as PWMs for stereotypical TFs and can improve performance for TFs with flexible binding characteristics. We present a new graphical representation of the motifs that convey properties of position interdependence. TFFMs have been assessed on ChIP-seq data sets coming from the ENCODE project, revealing that they can perform better than both PWMs and the dinucleotide weight matrix extension in discriminating ChIP-seq from background sequences. Under the assumption that ChIP-seq signal values are correlated with the affinity of the TF-DNA binding, we find that TFFM scores correlate with ChIP-seq peak signals. Moreover, using available TF-DNA affinity measurements for the Max TF, we demonstrate that TFFMs constructed from ChIP-seq data correlate with published experimentally measured DNA-binding affinities. Finally, TFFMs allow for the straightforward computation of an integrated TF occupancy score across a sequence. These results demonstrate the capacity of TFFMs to accurately model DNA-protein interactions, while providing a single unified framework suitable for the next generation of TFBS prediction.
Author Summary
Transcription factors are critical proteins for sequence-specific control of transcriptional regulation. Finding where these proteins bind to DNA is of key importance for global efforts to decipher the complex mechanisms of gene regulation. Greater understanding of the regulation of transcription promises to improve human genetic analysis by specifying critical gene components that have eluded investigators. Classically, computational prediction of transcription factor binding sites (TFBS) is based on models giving weights to each nucleotide at each position. We introduce a novel statistical model for the prediction of TFBS tolerant of a broader range of TFBS configurations than can be conveniently accommodated by existing methods. The new models are designed to address the confounding properties of nucleotide composition, inter-positional sequence dependence and variable lengths (e.g. variable spacing between half-sites) observed in the more comprehensive experimental data now emerging. The new models generate scores consistent with DNA-protein affinities measured experimentally and can be represented graphically, retaining desirable attributes of past methods. It demonstrates the capacity of the new approach to accurately assess DNA-protein interactions. With the rich experimental data generated from chromatin immunoprecipitation experiments, a greater diversity of TFBS properties has emerged that can now be accommodated within a single predictive approach.
doi:10.1371/journal.pcbi.1003214
PMCID: PMC3764009  PMID: 24039567
2.  LASAGNA: A novel algorithm for transcription factor binding site alignment 
BMC Bioinformatics  2013;14:108.
Background
Scientists routinely scan DNA sequences for transcription factor (TF) binding sites (TFBSs). Most of the available tools rely on position-specific scoring matrices (PSSMs) constructed from aligned binding sites. Because of the resolutions of assays used to obtain TFBSs, databases such as TRANSFAC, ORegAnno and PAZAR store unaligned variable-length DNA segments containing binding sites of a TF. These DNA segments need to be aligned to build a PSSM. While the TRANSFAC database provides scoring matrices for TFs, nearly 78% of the TFs in the public release do not have matrices available. As work on TFBS alignment algorithms has been limited, it is highly desirable to have an alignment algorithm tailored to TFBSs.
Results
We designed a novel algorithm named LASAGNA, which is aware of the lengths of input TFBSs and utilizes position dependence. Results on 189 TFs of 5 species in the TRANSFAC database showed that our method significantly outperformed ClustalW2 and MEME. We further compared a PSSM method dependent on LASAGNA to an alignment-free TFBS search method. Results on 89 TFs whose binding sites can be located in genomes showed that our method is significantly more precise at fixed recall rates. Finally, we described LASAGNA-ChIP, a more sophisticated version for ChIP (Chromatin immunoprecipitation) experiments. Under the one-per-sequence model, it showed comparable performance with MEME in discovering motifs in ChIP-seq peak sequences.
Conclusions
We conclude that the LASAGNA algorithm is simple and effective in aligning variable-length binding sites. It has been integrated into a user-friendly webtool for TFBS search and visualization called LASAGNA-Search. The tool currently stores precomputed PSSM models for 189 TFs and 133 TFs built from TFBSs in the TRANSFAC Public database (release 7.0) and the ORegAnno database (08Nov10 dump), respectively. The webtool is available at http://biogrid.engr.uconn.edu/lasagna_search/.
doi:10.1186/1471-2105-14-108
PMCID: PMC3747862  PMID: 23522376
3.  Tree-Based Position Weight Matrix Approach to Model Transcription Factor Binding Site Profiles 
PLoS ONE  2011;6(9):e24210.
Most of the position weight matrix (PWM) based bioinformatics methods developed to predict transcription factor binding sites (TFBS) assume each nucleotide in the sequence motif contributes independently to the interaction between protein and DNA sequence, usually producing high false positive predictions. The increasing availability of TF enrichment profiles from recent ChIP-Seq methodology facilitates the investigation of dependent structure and accurate prediction of TFBSs. We develop a novel Tree-based PWM (TPWM) approach to accurately model the interaction between TF and its binding site. The whole tree-structured PWM could be considered as a mixture of different conditional-PWMs. We propose a discriminative approach, called TPD (TPWM based Discriminative Approach), to construct the TPWM from the ChIP-Seq data with a pre-existing PWM. To achieve the maximum discriminative power between the positive and negative datasets, the cutoff value is determined based on the Matthew Correlation Coefficient (MCC). The resulting TPWMs are evaluated with respect to accuracy on extensive synthetic datasets. We then apply our TPWM discriminative approach on several real ChIP-Seq datasets to refine the current TFBS models stored in the TRANSFAC database. Experiments on both the simulated and real ChIP-Seq data show that the proposed method starting from existing PWM has consistently better performance than existing tools in detecting the TFBSs. The improved accuracy is the result of modelling the complete dependent structure of the motifs and better prediction of true positive rate. The findings could lead to better understanding of the mechanisms of TF-DNA interactions.
doi:10.1371/journal.pone.0024210
PMCID: PMC3166302  PMID: 21912677
4.  A Bayesian Search for Transcriptional Motifs 
PLoS ONE  2010;5(11):e13897.
Identifying transcription factor (TF) binding sites (TFBSs) is an important step towards understanding transcriptional regulation. A common approach is to use gaplessly aligned, experimentally supported TFBSs for a particular TF, and algorithmically search for more occurrences of the same TFBSs. The largest publicly available databases of TF binding specificities contain models which are represented as position weight matrices (PWM). There are other methods using more sophisticated representations, but these have more limited databases, or aren't publicly available. Therefore, this paper focuses on methods that search using one PWM per TF. An algorithm, MATCHTM, for identifying TFBSs corresponding to a particular PWM is available, but is not based on a rigorous statistical model of TF binding, making it difficult to interpret or adjust the parameters and output of the algorithm. Furthermore, there is no public description of the algorithm sufficient to exactly reproduce it. Another algorithm, MAST, computes a p-value for the presence of a TFBS using true probabilities of finding each base at each offset from that position. We developed a statistical model, BaSeTraM, for the binding of TFs to TFBSs, taking into account random variation in the base present at each position within a TFBS. Treating the counts in the matrices and the sequences of sites as random variables, we combine this TFBS composition model with a background model to obtain a Bayesian classifier. We implemented our classifier in a package (SBaSeTraM). We tested SBaSeTraM against a MATCHTM implementation by searching all probes used in an experimental Saccharomyces cerevisiae TF binding dataset, and comparing our predictions to the data. We found no statistically significant differences in sensitivity between the algorithms (at fixed selectivity), indicating that SBaSeTraM's performance is at least comparable to the leading currently available algorithm. Our software is freely available at: http://wiki.github.com/A1kmm/sbasetram/building-the-tools.
doi:10.1371/journal.pone.0013897
PMCID: PMC2987817  PMID: 21124986
5.  Pinpointing transcription factor binding sites from ChIP-seq data with SeqSite 
BMC Systems Biology  2011;5(Suppl 2):S3.
Background
Chromatin immunoprecipitation combined with the next-generation DNA sequencing technologies (ChIP-seq) becomes a key approach for detecting genome-wide sets of genomic sites bound by proteins, such as transcription factors (TFs). Several methods and open-source tools have been developed to analyze ChIP-seq data. However, most of them are designed for detecting TF binding regions instead of accurately locating transcription factor binding sites (TFBSs). It is still challenging to pinpoint TFBSs directly from ChIP-seq data, especially in regions with closely spaced binding events.
Results
With the aim to pinpoint TFBSs at a high resolution, we propose a novel method named SeqSite, implementing a two-step strategy: detecting tag-enriched regions first and pinpointing binding sites in the detected regions. The second step is done by modeling the tag density profile, locating TFBSs on each strand with a least-squares model fitting strategy, and merging the detections from the two strands. Experiments on simulation data show that SeqSite can locate most of the binding sites more than 40-bp from each other. Applications on three human TF ChIP-seq datasets demonstrate the advantage of SeqSite for its higher resolution in pinpointing binding sites compared with existing methods.
Conclusions
We have developed a computational tool named SeqSite, which can pinpoint both closely spaced and isolated binding sites, and consequently improves the resolution of TFBS detection from ChIP-seq data.
doi:10.1186/1752-0509-5-S2-S3
PMCID: PMC3287483  PMID: 22784574
6.  Global Mapping of Transcription Factor Binding Sites by Sequencing Chromatin Surrogates: a Perspective on Experimental Design, Data Analysis, and Open Problems 
Statistics in Biosciences  2012;5(1):156-178.
Mapping genome-wide binding sites of all transcription factors (TFs) in all biological contexts is a critical step toward understanding gene regulation. The state-of-the-art technologies for mapping transcription factor binding sites (TFBSs) couple chromatin immunoprecipitation (ChIP) with high-throughput sequencing (ChIP-seq) or tiling array hybridization (ChIP-chip). These technologies have limitations: they are low-throughput with respect to surveying many TFs. Recent advances in genome-wide chromatin profiling, including development of technologies such as DNase-seq, FAIRE-seq and ChIP-seq for histone modifications, make it possible to predict in vivo TFBSs by analyzing chromatin features at computationally determined DNA motif sites. This promising new approach may allow researchers to monitor the genome-wide binding sites of many TFs simultaneously. In this article, we discuss various experimental design and data analysis issues that arise when applying this approach. Through a systematic analysis of the data from the Encyclopedia Of DNA Elements (ENCODE) project, we compare the predictive power of individual and combinations of chromatin marks using supervised and unsupervised learning methods, and evaluate the value of integrating information from public ChIP and gene expression data. We also highlight the challenges and opportunities for developing novel analytical methods, such as resolving the one-motif-multiple-TF ambiguity and distinguishing functional and non-functional TF binding targets from the predicted binding sites.
Electronic Supplementary Material
The online version of this article (doi:10.1007/s12561-012-9066-5) contains supplementary material, which is available to authorized users.
doi:10.1007/s12561-012-9066-5
PMCID: PMC3677239  PMID: 23762209
Transcription factor binding sites; DNase-seq; ChIP-seq; FAIRE-seq; Next-generation sequencing; Motif
7.  MYBS: a comprehensive web server for mining transcription factor binding sites in yeast 
Nucleic Acids Research  2007;35(Web Server issue):W221-W226.
Correct interactions between transcription factors (TFs) and their binding sites (TFBSs) are of central importance to gene regulation. Recently developed chromatin-immunoprecipitation DNA chip (ChIP-chip) techniques and the phylogenetic footprinting method provide ways to identify TFBSs with high precision. In this study, we constructed a user-friendly interactive platform for dynamic binding site mapping using ChIP-chip data and phylogenetic footprinting as two filters. MYBS (Mining Yeast Binding Sites) is a comprehensive web server that integrates an array of both experimentally verified and predicted position weight matrixes (PWMs) from eleven databases, including 481 binding motif consensus sequences and 71 PWMs that correspond to 183 TFs. MYBS users can search within this platform for motif occurrences (possible binding sites) in the promoters of genes of interest via simple motif or gene queries in conjunction with the above two filters. In addition, MYBS enables users to visualize in parallel the potential regulators for a given set of genes, a feature useful for finding potential regulatory associations between TFs. MYBS also allows users to identify target gene sets of each TF pair, which could be used as a starting point for further explorations of TF combinatorial regulation. MYBS is available at http://cg1.iis.sinica.edu.tw/~mybs/.
doi:10.1093/nar/gkm379
PMCID: PMC1933147  PMID: 17537814
8.  Identification of co-occurring transcription factor binding sites from DNA sequence using clustered position weight matrices 
Nucleic Acids Research  2011;40(5):e38.
Accurate prediction of transcription factor binding sites (TFBSs) is a prerequisite for identifying cis-regulatory modules that underlie transcriptional regulatory circuits encoded in the genome. Here, we present a computational framework for detecting TFBSs, when multiple position weight matrices (PWMs) for a transcription factor are available. Grouping multiple PWMs of a transcription factor (TF) based on their sequence similarity improves the specificity of TFBS prediction, which was evaluated using multiple genome-wide ChIP-Seq data sets from 26 TFs. The Z-scores of the area under a receiver operating characteristic curve (AUC) values of 368 TFs were calculated and used to statistically identify co-occurring regulatory motifs in the TF bound ChIP loci. Motifs that are co-occurring along with the empirical bindings of E2F, JUN or MYC have been evaluated, in the basal or stimulated condition. Results prove our method can be useful to systematically identify the co-occurring motifs of the TF for the given conditions.
doi:10.1093/nar/gkr1252
PMCID: PMC3300004  PMID: 22187154
9.  A Structural-Based Strategy for Recognition of Transcription Factor Binding Sites 
PLoS ONE  2013;8(1):e52460.
Scanning through genomes for potential transcription factor binding sites (TFBSs) is becoming increasingly important in this post-genomic era. The position weight matrix (PWM) is the standard representation of TFBSs utilized when scanning through sequences for potential binding sites. However, many transcription factor (TF) motifs are short and highly degenerate, and methods utilizing PWMs to scan for sites are plagued by false positives. Furthermore, many important TFs do not have well-characterized PWMs, making identification of potential binding sites even more difficult. One approach to the identification of sites for these TFs has been to use the 3D structure of the TF to predict the DNA structure around the TF and then to generate a PWM from the predicted 3D complex structure. However, this approach is dependent on the similarity of the predicted structure to the native structure. We introduce here a novel approach to identify TFBSs utilizing structure information that can be applied to TFs without characterized PWMs, as long as a 3D complex structure (TF/DNA) exists. This approach utilizes an energy function that is uniquely trained on each structure. Our approach leads to increased prediction accuracy and robustness compared with those using a more general energy function. The software is freely available upon request.
doi:10.1371/journal.pone.0052460
PMCID: PMC3540023  PMID: 23320072
10.  Dinucleotide Weight Matrices for Predicting Transcription Factor Binding Sites: Generalizing the Position Weight Matrix 
PLoS ONE  2010;5(3):e9722.
Background
Identifying transcription factor binding sites (TFBS) in silico is key in understanding gene regulation. TFBS are string patterns that exhibit some variability, commonly modelled as “position weight matrices” (PWMs). Though convenient, the PWM has significant limitations, in particular the assumed independence of positions within the binding motif; and predictions based on PWMs are usually not very specific to known functional sites. Analysis here on binding sites in yeast suggests that correlation of dinucleotides is not limited to near-neighbours, but can extend over considerable gaps.
Methodology/Principal Findings
I describe a straightforward generalization of the PWM model, that considers frequencies of dinucleotides instead of individual nucleotides. Unlike previous efforts, this method considers all dinucleotides within an extended binding region, and does not make an attempt to determine a priori the significance of particular dinucleotide correlations. I describe how to use a “dinucleotide weight matrix” (DWM) to predict binding sites, dealing in particular with the complication that its entries are not independent probabilities. Benchmarks show, for many factors, a dramatic improvement over PWMs in precision of predicting known targets. In most cases, significant further improvement arises by extending the commonly defined “core motifs” by about 10bp on either side. Though this flanking sequence shows no strong motif at the nucleotide level, the predictive power of the dinucleotide model suggests that the “signature” in DNA sequence of protein-binding affinity extends beyond the core protein-DNA contact region.
Conclusion/Significance
While computationally more demanding and slower than PWM-based approaches, this dinucleotide method is straightforward, both conceptually and in implementation, and can serve as a basis for future improvements.
doi:10.1371/journal.pone.0009722
PMCID: PMC2842295  PMID: 20339533
11.  PolyaPeak: Detecting Transcription Factor Binding Sites from ChIP-seq Using Peak Shape Information 
PLoS ONE  2014;9(3):e89694.
ChIP-seq is a powerful technology for detecting genomic regions where a protein of interest interacts with DNA. ChIP-seq data for mapping transcription factor binding sites (TFBSs) have a characteristic pattern: around each binding site, sequence reads aligned to the forward and reverse strands of the reference genome form two separate peaks shifted away from each other, and the true binding site is located in between these two peaks. While it has been shown previously that the accuracy and resolution of binding site detection can be improved by modeling the pattern, efficient methods are unavailable to fully utilize that information in TFBS detection procedure.
We present PolyaPeak, a new method to improve TFBS detection by incorporating the peak shape information. PolyaPeak describes peak shapes using a flexible Pólya model. The shapes are automatically learnt from the data using Minorization-Maximization (MM) algorithm, then integrated with the read count information via a hierarchical model to distinguish true binding sites from background noises.
Extensive real data analyses show that PolyaPeak is capable of robustly improving TFBS detection compared with existing methods. An R package is freely available.
doi:10.1371/journal.pone.0089694
PMCID: PMC3946423  PMID: 24608116
12.  Direct vs 2-stage approaches to structured motif finding 
Background
The notion of DNA motif is a mathematical abstraction used to model regions of the DNA (known as Transcription Factor Binding Sites, or TFBSs) that are bound by a given Transcription Factor to regulate gene expression or repression. In turn, DNA structured motifs are a mathematical counterpart that models sets of TFBSs that work in concert in the gene regulations processes of higher eukaryotic organisms. Typically, a structured motif is composed of an ordered set of isolated (or simple) motifs, separated by a variable, but somewhat constrained number of “irrelevant” base-pairs. Discovering structured motifs in a set of DNA sequences is a computationally hard problem that has been addressed by a number of authors using either a direct approach, or via the preliminary identification and successive combination of simple motifs.
Results
We describe a computational tool, named SISMA, for the de-novo discovery of structured motifs in a set of DNA sequences. SISMA is an exact, enumerative algorithm, meaning that it finds all the motifs conforming to the specifications. It does so in two stages: first it discovers all the possible component simple motifs, then combines them in a way that respects the given constraints. We developed SISMA mainly with the aim of understanding the potential benefits of such a 2-stage approach w.r.t. direct methods. In fact, no 2-stage software was available for the general problem of structured motif discovery, but only a few tools that solved restricted versions of the problem. We evaluated SISMA against other published tools on a comprehensive benchmark made of both synthetic and real biological datasets. In a significant number of cases, SISMA outperformed the competitors, exhibiting a good performance also in most of the cases in which it was inferior.
Conclusions
A reflection on the results obtained lead us to conclude that a 2-stage approach can be implemented with many advantages over direct approaches. Some of these have to do with greater modularity, ease of parallelization, and the possibility to perform adaptive searches of structured motifs. As another consideration, we noted that most hard instances for SISMA were easy to detect in advance. In these cases one may initially opt for a direct method; or, as a viable alternative in most laboratories, one could run both direct and 2-stage tools in parallel, halting the computations when the first halts.
doi:10.1186/1748-7188-7-20
PMCID: PMC3564690  PMID: 22908910
Structured motif; TFBS discovery; Combinatorial algorithms
13.  Optimized Position Weight Matrices in Prediction of Novel Putative Binding Sites for Transcription Factors in the Drosophila melanogaster Genome 
PLoS ONE  2013;8(8):e68712.
Position weight matrices (PWMs) have become a tool of choice for the identification of transcription factor binding sites in DNA sequences. DNA-binding proteins often show degeneracy in their binding requirement and thus the overall binding specificity of many proteins is unknown and remains an active area of research. Although existing PWMs are more reliable predictors than consensus string matching, they generally result in a high number of false positive hits. Our previous study introduced a promising approach to PWM refinement in which known motifs are used to computationally mine putative binding sites directly from aligned promoter regions using composition of similar sites. In the present study, we extended this technique originally tested on single examples of transcription factors (TFs) and showed its capability to optimize PWM performance to predict new binding sites in the fruit fly genome. We propose refined PWMs in mono- and dinucleotide versions similarly computed for a large variety of transcription factors of Drosophila melanogaster. Along with the addition of many auxiliary sites the optimization includes variation of the PWM motif length, the binding sites location on the promoters and the PWM score threshold. To assess the predictive performance of the refined PWMs we compared them to conventional TRANSFAC and JASPAR sources. The results have been verified using performed tests and literature review. Overall, the refined PWMs containing putative sites derived from real promoter content processed using optimized parameters had better general accuracy than conventional PWMs.
doi:10.1371/journal.pone.0068712
PMCID: PMC3735551  PMID: 23936309
14.  A flexible integrative approach based on random forest improves prediction of transcription factor binding sites 
Nucleic Acids Research  2012;40(14):e106.
Transcription factor binding sites (TFBSs) are DNA sequences of 6–15 base pairs. Interaction of these TFBSs with transcription factors (TFs) is largely responsible for most spatiotemporal gene expression patterns. Here, we evaluate to what extent sequence-based prediction of TFBSs can be improved by taking into account the positional dependencies of nucleotides (NPDs) and the nucleotide sequence-dependent structure of DNA. We make use of the random forest algorithm to flexibly exploit both types of information. Results in this study show that both the structural method and the NPD method can be valuable for the prediction of TFBSs. Moreover, their predictive values seem to be complementary, even to the widely used position weight matrix (PWM) method. This led us to combine all three methods. Results obtained for five eukaryotic TFs with different DNA-binding domains show that our method improves classification accuracy for all five eukaryotic TFs compared with other approaches. Additionally, we contrast the results of seven smaller prokaryotic sets with high-quality data and show that with the use of high-quality data we can significantly improve prediction performance. Models developed in this study can be of great use for gaining insight into the mechanisms of TF binding.
doi:10.1093/nar/gks283
PMCID: PMC3413102  PMID: 22492513
15.  Low nucleosome occupancy is encoded around functional human transcription factor binding sites 
BMC Genomics  2008;9:332.
Background
Transcriptional regulation of genes in eukaryotes is achieved by the interactions of multiple transcription factors with arrays of transcription factor binding sites (TFBSs) on DNA and with each other. Identification of these TFBSs is an essential step in our understanding of gene regulatory networks, but computational prediction of TFBSs with either consensus or commonly used stochastic models such as Position-Specific Scoring Matrices (PSSMs) results in an unacceptably high number of hits consisting of a few true functional binding sites and numerous false non-functional binding sites. This is due to the inability of the models to incorporate higher order properties of sequences including sequences surrounding TFBSs and influencing the positioning of nucleosomes and/or the interactions that might occur between transcription factors.
Results
Significant improvement can be expected through the development of a new framework for the modeling and prediction of TFBSs that considers explicitly these higher order sequence properties. It would be particularly interesting to include in the new modeling framework the information present in the nucleosome positioning sequences (NPSs) surrounding TFBSs, as it can be hypothesized that genomes use this information to encode the formation of stable nucleosomes over non-functional sites, while functional sites have a more open chromatin configuration.
In this report we evaluate the usefulness of the latter feature by comparing the nucleosome occupancy probabilities around experimentally verified human TFBSs with the nucleosome occupancy probabilities around false positive TFBSs and in random sequences.
Conclusion
We present evidence that nucleosome occupancy is remarkably lower around true functional human TFBSs as compared to non-functional human TFBSs, which supports the use of this feature to improve current TFBS prediction approaches in higher eukaryotes.
doi:10.1186/1471-2164-9-332
PMCID: PMC2490708  PMID: 18627598
16.  Genome-wide transcription factor binding: beyond direct target regulation 
Trends in genetics : TIG  2011;27(4):141-148.
The binding of transcription factors to specific DNA target sequences is the fundamental basis of gene regulatory networks. Chromatin immunoprecipitation combined with DNA tiling arrays or high-throughput sequencing—ChIP-chip and ChIP-seq—has produced many recent studies that detail the binding sites of various transcription factors. Surprisingly, data from a variety of model organisms and tissues have demonstrated that transcription factors vary greatly in their number of genomic binding sites, and that binding events can significantly exceed the number of known or possible direct gene targets. Thus, our current understanding of transcription factor function must expand to encompass what role, if any, binding might play outside of direct transcriptional target regulation. Here, we discuss the biological significance of genome-wide binding of transcription factors and present models that can account for this phenomenon.
doi:10.1016/j.tig.2011.01.001
PMCID: PMC3068217  PMID: 21295369
17.  ChIPBase: a database for decoding the transcriptional regulation of long non-coding RNA and microRNA genes from ChIP-Seq data 
Nucleic Acids Research  2012;41(D1):D177-D187.
Long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) represent two classes of important non-coding RNAs in eukaryotes. Although these non-coding RNAs have been implicated in organismal development and in various human diseases, surprisingly little is known about their transcriptional regulation. Recent advances in chromatin immunoprecipitation with next-generation DNA sequencing (ChIP-Seq) have provided methods of detecting transcription factor binding sites (TFBSs) with unprecedented sensitivity. In this study, we describe ChIPBase (http://deepbase.sysu.edu.cn/chipbase/), a novel database that we have developed to facilitate the comprehensive annotation and discovery of transcription factor binding maps and transcriptional regulatory relationships of lncRNAs and miRNAs from ChIP-Seq data. The current release of ChIPBase includes high-throughput sequencing data that were generated by 543 ChIP-Seq experiments in diverse tissues and cell lines from six organisms. By analysing millions of TFBSs, we identified tens of thousands of TF-lncRNA and TF-miRNA regulatory relationships. Furthermore, two web-based servers were developed to annotate and discover transcriptional regulatory relationships of lncRNAs and miRNAs from ChIP-Seq data. In addition, we developed two genome browsers, deepView and genomeView, to provide integrated views of multidimensional data. Moreover, our web implementation supports diverse query types and the exploration of TFs, lncRNAs, miRNAs, gene ontologies and pathways.
doi:10.1093/nar/gks1060
PMCID: PMC3531181  PMID: 23161675
18.  Frequent Gain and Loss of Functional Transcription Factor Binding Sites 
PLoS Computational Biology  2007;3(5):e99.
Cis-regulatory sequences are not always conserved across species. Divergence within cis-regulatory sequences may result from the evolution of species-specific patterns of gene expression or the flexible nature of the cis-regulatory code. The identification of functional divergence in cis-regulatory sequences is therefore important for both understanding the role of gene regulation in evolution and annotating regulatory elements. We have developed an evolutionary model to detect the loss of constraint on individual transcription factor binding sites (TFBSs). We find that a significant fraction of functionally constrained binding sites have been lost in a lineage-specific manner among three closely related yeast species. Binding site loss has previously been explained by turnover, where the concurrent gain and loss of a binding site maintains gene regulation. We estimate that nearly half of all loss events cannot be explained by binding site turnover. Recreating the mutations that led to binding site loss confirms that these sequence changes affect gene expression in some cases. We also estimate that there is a high rate of binding site gain, as more than half of experimentally identified S. cerevisiae binding sites are not conserved across species. The frequent gain and loss of TFBSs implies that cis-regulatory sequences are labile and, in the absence of turnover, may contribute to species-specific patterns of gene expression.
Author Summary
Research in the field of molecular evolution is focused on understanding the genetic basis of functional differences between species. Protein coding sequences have traditionally been the focus of these studies, as the genetic code enables a detailed study of the strength of selection acting on amino acid sequences. However, from the earliest cross-species sequence comparisons, it was clear that protein sequences among closely related species are too similar to explain the observed phenotypic diversity. This led to the hypothesis that the evolution of gene regulation has played a key role in generating diversity between species. The availability of numerous complete genome sequences has made it possible to begin testing this hypothesis. In this work, the authors use an evolutionary model to identify functional divergence within transcription factor binding sites, the core functional elements involved in gene regulation. Applying this model to the baker's yeast, Saccharomyces cerevisiae, and its three closest relatives, the authors find that a substantial fraction of the ancestral binding sites have been lost in a species-specific manner. In some cases the loss of the binding site creates gene expression differences that may be indicative of species-specific changes in gene regulation. This work provides a useful computational framework that will allow further study of the conservation of cis-regulatory sequences and their role in molecular evolution.
doi:10.1371/journal.pcbi.0030099
PMCID: PMC1876492  PMID: 17530920
19.  3DTF: a web server for predicting transcription factor PWMs using 3D structure-based energy calculations 
Nucleic Acids Research  2012;40(Web Server issue):W180-W185.
We present the webserver 3D transcription factor (3DTF) to compute position-specific weight matrices (PWMs) of transcription factors using a knowledge-based statistical potential derived from crystallographic data on protein–DNA complexes. Analysis of available structures that can be used to construct PWMs shows that there are hundreds of 3D structures from which PWMs could be derived, as well as thousands of proteins homologous to these. Therefore, we created 3DTF, which delivers binding matrices given the experimental or modeled protein–DNA complex. The webserver can be used by biologists to derive novel PWMs for transcription factors lacking known binding sites and is freely accessible at http://www.gene-regulation.com/pub/programs/3dtf/.
doi:10.1093/nar/gks551
PMCID: PMC3394331  PMID: 22693215
20.  Computational technique for improvement of the position-weight matrices for the DNA/protein binding sites 
Nucleic Acids Research  2005;33(7):2290-2301.
Position-weight matrices (PWMs) are broadly used to locate transcription factor binding sites in DNA sequences. The majority of existing PWMs provide a low level of both sensitivity and specificity. We present a new computational algorithm, a modification of the Staden–Bucher approach, that improves the PWM. We applied the proposed technique on the PWM of the GC-box, binding site for Sp1. The comparison of old and new PWMs shows that the latter increase both sensitivity and specificity. The statistical parameters of GC-box distribution in promoter regions and in the human genome, as well as in each chromosome, are presented. The majority of commonly used PWMs are the 4-row mononucleotide matrices, although 16-row dinucleotide matrices are known to be more informative. The algorithm efficiently determines the 16-row matrices and preliminary results show that such matrices provide better results than 4-row matrices.
doi:10.1093/nar/gki519
PMCID: PMC1084321  PMID: 15849315
21.  Creating PWMs of transcription factors using 3D structure-based computation of protein-DNA free binding energies 
BMC Bioinformatics  2010;11:225.
Background
Knowledge of transcription factor-DNA binding patterns is crucial for understanding gene transcription. Numerous DNA-binding proteins are annotated as transcription factors in the literature, however, for many of them the corresponding DNA-binding motifs remain uncharacterized.
Results
The position weight matrices (PWMs) of transcription factors from different structural classes have been determined using a knowledge-based statistical potential. The scoring function calibrated against crystallographic data on protein-DNA contacts recovered PWMs of various members of widely studied transcription factor families such as p53 and NF-κB. Where it was possible, extensive comparison to experimental binding affinity data and other physical models was made. Although the p50p50, p50RelB, and p50p65 dimers belong to the same family, particular differences in their PWMs were detected, thereby suggesting possibly different in vivo binding modes. The PWMs of p63 and p73 were computed on the basis of homology modeling and their performance was studied using upstream sequences of 85 p53/p73-regulated human genes. Interestingly, about half of the p63 and p73 hits reported by the Match algorithm in the altogether 126 promoters lay more than 2 kb upstream of the corresponding transcription start sites, which deviates from the common assumption that most regulatory sites are located more proximal to the TSS. The fact that in most of the cases the binding sites of p63 and p73 did not overlap with the p53 sites suggests that p63 and p73 could influence the p53 transcriptional activity cooperatively. The newly computed p50p50 PWM recovered 5 more experimental binding sites than the corresponding TRANSFAC matrix, while both PWMs showed comparable receiver operator characteristics.
Conclusions
A novel algorithm was developed to calculate position weight matrices from protein-DNA complex structures. The proposed algorithm was extensively validated against experimental data. The method was further combined with Homology Modeling to obtain PWMs of factors for which crystallographic complexes with DNA are not yet available. The performance of PWMs obtained in this work in comparison to traditionally constructed matrices demonstrates that the structure-based approach presents a promising alternative to experimental determination of transcription factor binding properties.
doi:10.1186/1471-2105-11-225
PMCID: PMC2879287  PMID: 20438625
22.  Improved predictions of transcription factor binding sites using physicochemical features of DNA 
Nucleic Acids Research  2012;40(22):e175.
Typical approaches for predicting transcription factor binding sites (TFBSs) involve use of a position-specific weight matrix (PWM) to statistically characterize the sequences of the known sites. Recently, an alternative physicochemical approach, called SiteSleuth, was proposed. In this approach, a linear support vector machine (SVM) classifier is trained to distinguish TFBSs from background sequences based on local chemical and structural features of DNA. SiteSleuth appears to generally perform better than PWM-based methods. Here, we improve the SiteSleuth approach by considering both new physicochemical features and algorithmic modifications. New features are derived from Gibbs energies of amino acid–DNA interactions and hydroxyl radical cleavage profiles of DNA. Algorithmic modifications consist of inclusion of a feature selection step, use of a nonlinear kernel in the SVM classifier, and use of a consensus-based post-processing step for predictions. We also considered SVM classification based on letter features alone to distinguish performance gains from use of SVM-based models versus use of physicochemical features. The accuracy of each of the variant methods considered was assessed by cross validation using data available in the RegulonDB database for 54 Escherichia coli TFs, as well as by experimental validation using published ChIP-chip data available for Fis and Lrp.
doi:10.1093/nar/gks771
PMCID: PMC3526315  PMID: 22923524
23.  Increasing Coverage of Transcription Factor Position Weight Matrices through Domain-level Homology 
PLoS ONE  2012;7(8):e42779.
Transcription factor-DNA interactions, central to cellular regulation and control, are commonly described by position weight matrices (PWMs). These matrices are frequently used to predict transcription factor binding sites in regulatory regions of DNA to complement and guide further experimental investigation. The DNA sequence preferences of transcription factors, encoded in PWMs, are dictated primarily by select residues within the DNA binding domain(s) that interact directly with DNA. Therefore, the DNA binding properties of homologous transcription factors with identical DNA binding domains may be characterized by PWMs derived from different species. Accordingly, we have implemented a fully automated domain-level homology searching method for identical DNA binding sequences.
By applying the domain-level homology search to transcription factors with existing PWMs in the JASPAR and TRANSFAC databases, we were able to significantly increase coverage in terms of the total number of PWMs associated with a given species, assign PWMs to transcription factors that did not previously have any associations, and increase the number of represented species with PWMs over an order of magnitude. Additionally, using protein binding microarray (PBM) data, we have validated the domain-level method by demonstrating that transcription factor pairs with matching DNA binding domains exhibit comparable DNA binding specificity predictions to transcription factor pairs with completely identical sequences.
The increased coverage achieved herein demonstrates the potential for more thorough species-associated investigation of protein-DNA interactions using existing resources. The PWM scanning results highlight the challenging nature of transcription factors that contain multiple DNA binding domains, as well as the impact of motif discovery on the ability to predict DNA binding properties. The method is additionally suitable for identifying domain-level homology mappings to enable utilization of additional information sources in the study of transcription factors. The domain-level homology search method, resulting PWM mappings, web-based user interface, and web API are publicly available at http://dodoma.systemsbiology.netdodoma.systemsbiology.net.
doi:10.1371/journal.pone.0042779
PMCID: PMC3428306  PMID: 22952610
24.  An information transmission model for transcription factor binding at regulatory DNA sites 
Background
Computational identification of transcription factor binding sites (TFBSs) is a rapid, cost-efficient way to locate unknown regulatory elements. With increased potential for high-throughput genome sequencing, the availability of accurate computational methods for TFBS prediction has never been as important as it currently is. To date, identifying TFBSs with high sensitivity and specificity is still an open challenge, necessitating the development of novel models for predicting transcription factor-binding regulatory DNA elements.
Results
Based on the information theory, we propose a model for transcription factor binding of regulatory DNA sites. Our model incorporates position interdependencies in effective ways. The model computes the information transferred (TI) between the transcription factor and the TFBS during the binding process and uses TI as the criterion to determine whether the sequence motif is a possible TFBS. Based on this model, we developed a computational method to identify TFBSs. By theoretically proving and testing our model using both real and artificial data, we found that our model provides highly accurate predictive results.
Conclusions
In this study, we present a novel model for transcription factor binding regulatory DNA sites. The model can provide an increased ability to detect TFBSs.
doi:10.1186/1742-4682-9-19
PMCID: PMC3442977  PMID: 22672438
25.  Improved benchmarks for computational motif discovery 
BMC Bioinformatics  2007;8:193.
Background
An important step in annotation of sequenced genomes is the identification of transcription factor binding sites. More than a hundred different computational methods have been proposed, and it is difficult to make an informed choice. Therefore, robust assessment of motif discovery methods becomes important, both for validation of existing tools and for identification of promising directions for future research.
Results
We use a machine learning perspective to analyze collections of transcription factors with known binding sites. Algorithms are presented for finding position weight matrices (PWMs), IUPAC-type motifs and mismatch motifs with optimal discrimination of binding sites from remaining sequence. We show that for many data sets in a recently proposed benchmark suite for motif discovery, none of the common motif models can accurately discriminate the binding sites from remaining sequence. This may obscure the distinction between the potential performance of the motif discovery tool itself versus the intrinsic complexity of the problem we are trying to solve. Synthetic data sets may avoid this problem, but we show on some previously proposed benchmarks that there may be a strong bias towards a presupposed motif model. We also propose a new approach to benchmark data set construction. This approach is based on collections of binding site fragments that are ranked according to the optimal level of discrimination achieved with our algorithms. This allows us to select subsets with specific properties. We present one benchmark suite with data sets that allow good discrimination between positive and negative instances with the common motif models. These data sets are suitable for evaluating algorithms for motif discovery that rely on these models. We present another benchmark suite where PWM, IUPAC and mismatch motif models are not able to discriminate reliably between positive and negative instances. This suite could be used for evaluating more powerful motif models.
Conclusion
Our improved benchmark suites have been designed to differentiate between the performance of motif discovery algorithms and the power of motif models. We provide a web server where users can download our benchmark suites, submit predictions and visualize scores on the benchmarks.
doi:10.1186/1471-2105-8-193
PMCID: PMC1903367  PMID: 17559676

Results 1-25 (1050188)