ChIP-exo signal associated with DNA-binding motifs provides insight into the genomic binding of the glucocorticoid receptor and cooperating transcription factors.

Stephan R Starick,Michael I Love,Sebastiaan H Meijsing,Jonas Ibn-Salem,Marcel Jurk,Martin Vingron,Ho-Ryun Chung,Céline Hernandez,Morgane Thomas-Chollier

doi:10.1101/gr.185157.114

Abstract

The classical DNA recognition sequence of the glucocorticoid receptor (GR) appears to be present at only a fraction of bound genomic regions. To identify sequences responsible for recruitment of this transcription factor (TF) to individual loci, we turned to the high-resolution ChIP-exo approach. We exploited this signal by determining footprint profiles of TF binding at single-base-pair resolution using ExoProfiler, a computational pipeline based on DNA binding motifs. When applied to our GR and the few available public ChIP-exo data sets, we find that ChIP-exo footprints are protein- and recognition sequence-specific signatures of genomic TF association. Furthermore, we show that ChIP-exo captures information about TFs other than the one directly targeted by the antibody in the ChIP procedure. Consequently, the shape of the ChIP-exo footprint can be used to discriminate between direct and indirect (tethering to other DNA-bound proteins) DNA association of GR. Together, our findings indicate that the absence of classical recognition sequences can be explained by direct GR binding to a broader spectrum of sequences than previously known, either as a homodimer or as a heterodimer binding together with a member of the ETS or TEAD families of TFs, or alternatively by indirect recruitment via FOX or STAT proteins. ChIP-exo footprints also bring structural insights and locate DNA:protein cross-link points that are compatible with crystal structures of the studied TFs. Overall, our generically applicable footprint-based approach uncovers new structural and functional insights into the diverse ways of genomic cooperation and association of TFs.

Highlights

Transcriptional regulatory factors (TFs) control where, when, and at which level a gene is expressed by binding to specific regulatory sequences associated with their target genes
If and how such predicted sites are bound by the TF of interest is unclear for several reasons: (1) enriched sequences do not necessarily indicate TF binding; (2) they can be directly recognized by the TF studied or be involved in tethering it to the DNA; and (3) they might recruit other TFs that play a role in opening specific genomic regions and are not directly involved in providing a physical connection between DNA and the studied TF
Because ChIP-seq does not have the resolution to discriminate between these scenarios, we turned to ChIP-exo (Rhee and Pugh 2011) to study the genomic interactions of glucocorticoid receptor (GR) at higher resolution

Summary

Introduction

Transcriptional regulatory factors (TFs) control where, when, and at which level a gene is expressed by binding to specific regulatory sequences associated with their target genes. Candidates for TF binding sites (TFBSs) result from computational analyses aimed at identifying overrepresented sequence motifs within bound regions. If and how such predicted sites are bound by the TF of interest is unclear for several reasons: (1) enriched sequences do not necessarily indicate TF binding; (2) they can be directly recognized by the TF studied or be involved in tethering it to the DNA; and (3) they might recruit other TFs that play a role in opening specific genomic regions and are not directly involved in providing a physical connection between DNA and the studied TF. These limitations are, in part, a consequence of the limited resolution of ChIP-seq that identifies regions rather than the exact location of TF binding. The resulting footprint profiles enabled us to generate new testable hypotheses, leading to new insights into mechanisms responsible for recruiting GR to specific genomic loci

Methods

Results

Conclusion