Dataset Information

Direct ChIP-Seq significance analysis improves target prediction.

ABSTRACT: Chromatin immunoprecipitation followed by sequencing of protein-bound DNA fragments (ChIP-Seq) is an effective high-throughput methodology for the identification of context specific DNA fragments that are bound by specific proteins in vivo. Despite significant progress in the bioinformatics analysis of this genome-scale data, a number of challenges remain as technology-dependent biases, including variable target accessibility and mappability, sequence-dependent variability, and non-specific binding affinity must be accounted for.We introduce a nonparametric method for scoring consensus regions of aligned immunoprecipitated DNA fragments when appropriate control experiments are available. Our method uses local models for null binding; these are necessary because binding prediction scores based on global models alone fail to properly account for specialized features of genomic regions and chance pull downs of specific DNA fragments, thus disproportionally rewarding some genomic regions and decreasing prediction accuracy. We make no assumptions about the structure or amplitude of bound peaks, yet we show that our method outperforms leading methods developed using either global or local null hypothesis models for random binding. We test prediction performance by comparing analyses of ChIP-seq, ChIP-chip, motif-based binding-site prediction, and shRNA assays, showing high reproducibility, binding-site enrichment in predicted target regions, and functional regulation of predicted targets.Given appropriate controls, a direct nonparametric method for identifying transcription-factor targets from ChIP-Seq assays may lead to both higher sensitivity and higher specificity, and should be preferred or used in conjunction with methods that use parametric models for null binding.

SUBMITTER: Bansal M

PROVIDER: S-EPMC4460594 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Direct ChIP-Seq significance analysis improves target prediction.

Bansal Mukesh M Mendiratta Geetu G Anand Santosh S Kushwaha Ritu R Kim Ryan R Kustagi Manju M Iyer Archana A Chaganti Raju S K RS Califano Andrea A Sumazin Pavel P

BMC genomics 20150526

<h4>Background</h4>Chromatin immunoprecipitation followed by sequencing of protein-bound DNA fragments (ChIP-Seq) is an effective high-throughput methodology for the identification of context specific DNA fragments that are bound by specific proteins in vivo. Despite significant progress in the bioinformatics analysis of this genome-scale data, a number of challenges remain as technology-dependent biases, including variable target accessibility and mappability, sequence-dependent variability, an ...[more]

PMID: 26040656

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Project description:Cardiac fibrosis is a common pathological feature in cardiac remodeling. This study aimed to explore the role of KDM5A in cardiac fibrosis via bioinformatics analysis. Cardiac fibroblasts (CFs) were harvested and cultured from 10 dilated cardiomyopathy (DCM) patients who underwent heart transplantation. Western blotting was applied to verify that KDM5A is regulated by angiotensin II (Ang II) via the PI3k/AKT signaling pathway. The differentially expressed genes (DEGs) were analyzed by transcriptomics. ChIP-seq and ChIP-qPCR were used to identify the genes bound by KDM5A. In integrative analysis, weighted gene coexpression network analysis (WGCNA) was performed to identify highly relevant gene modules. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed for the key genes in modules. The STRING database, Cytoscape, and MCODE were applied to construct the protein-protein interaction (PPI) network and screen hub genes. To verify the expression of DEGs regulated by KDM5A, Western blotting and immunofluorescence were performed in myocardial tissue samples. Immunofluorescence verified the vimentin positivity of CFs. Ang II upregulated the expression of KDM5A in CFs via the PI3K/AKT signaling pathway. GO analysis of DEGs indicated that regulation of vasoconstriction, extracellular region, and calcium ion binding were enriched when KDM5A interfered with CPI or Ang II. KEGG analysis of the DEGs revealed the involvement of ECM-receptor interaction, focal adhesion, PI3K-Akt signaling pathway, cell adhesion, and arrhythmogenic right ventricular cardiomyopathy pathways. Three hub genes (IGF1, MYH11, and TGFB3) were identified via four different algorithms. Subsequent verification in patient samples demonstrated that the hub genes, which were regulated by KDM5A, were downregulated in DCM samples. KDM5A is a key regulator in the progression of cardiac fibrosis. In this successful integrative analysis, IGF1, MYH11, and TGFB3 were determined to be coordinately expressed to participate in cardiac fibrosis.

Project description:BackgroundChIPx (i.e., ChIP-seq and ChIP-chip) is increasingly used to map genome-wide transcription factor (TF) binding sites. A single ChIPx experiment can identify thousands of TF bound genes, but typically only a fraction of these genes are functional targets that respond transcriptionally to perturbations of TF expression. To identify promising functional target genes for follow-up studies, researchers usually collect gene expression data from TF perturbation experiments to determine which of the TF targets respond transcriptionally to binding. Unfortunately, approximately 40% of ChIPx studies do not have accompanying gene expression data from TF perturbation experiments. For these studies, genes are often prioritized solely based on the binding strengths of ChIPx signals in order to choose follow-up candidates. ChIPXpress is a novel method that improves upon this ChIPx-only ranking approach by integrating ChIPx data with large amounts of Publicly available gene Expression Data (PED).ResultsWe demonstrate that PED does contain useful information to identify functional TF target genes despite its inherent heterogeneity. A truncated absolute correlation measure is developed to better capture the regulatory relationships between TFs and their target genes in PED. By integrating the information from ChIPx and PED, ChIPXpress can significantly increase the chance of finding functional target genes responsive to TF perturbation among the top ranked genes. ChIPXpress is implemented as an easy-to-use R/Bioconductor package. We evaluate ChIPXpress using 10 different ChIPx datasets in mouse and human and find that ChIPXpress rankings are more accurate than rankings based solely on ChIPx data and may result in substantial improvement in prediction accuracy, irrespective of which peak calling algorithm is used to analyze the ChIPx data.ConclusionsChIPXpress provides a new tool to better prioritize TF bound genes from ChIPx experiments for follow-up studies when investigators do not have their own gene expression data. It demonstrates that the regulatory information from PED can be used to boost ChIPx data analyses. It also represents an important step towards more fully utilizing the valuable, but highly heterogeneous data contained in public gene expression databases.

Dataset Information

Direct ChIP-Seq significance analysis improves target prediction.

Publications

Direct ChIP-Seq significance analysis improves target prediction.

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