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High-throughput sequencing identifies STAT3 as the DNA-associated factor for p53-NF-κB-complex-dependent gene expression in human heart failure

Mun-Kit Choy1, Mehregan Movassagh1, Lee Siggens1, Ana Vujic1, Martin Goddard2, Ana Sánchez3, Neil Perkins3, Nichola Figg1, Martin Bennett1, Jason Carroll4 and Roger Foo1*

Author Affiliations

1 Department of Medicine, University of Cambridge, Addenbrooke's Centre for Clinical Investigation, Hills Road, Cambridge, CB2 0QQ, UK

2 Department of Histopathology, Papworth Hospital, Papworth Everard, Cambridge, CB23 3RE, UK

3 Department of Cellular and Molecular Medicine, University of Bristol, School of Medical Sciences, University Walk, Bristol, BS8 1TD, UK

4 Cancer Research UK, Cambridge Research Institute, Li Ka Shing Centre, Robinson Way, Cambridge CB2 0RE, UK

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Genome Medicine 2010, 2:37  doi:10.1186/gm158

Published: 14 June 2010

Abstract

Background

Genome-wide maps of DNA regulatory elements and their interaction with transcription factors may form a framework for understanding regulatory circuits and gene expression control in human disease, but how these networks, comprising transcription factors and DNA-binding proteins, form complexes, interact with DNA and modulate gene expression remains largely unknown.

Methods

Using microRNA-21 (mir-21), which is an example of genes that are regulated in heart failure, we performed chromatin immunoprecipitation (ChIP) assays to determine the occupancy of transcription factors at this genetic locus. Tissue ChIP was further performed using human hearts and genome-wide occupancies of these transcription factors were analyzed by high-throughput sequencing.

Results

We show that the transcription factor p53 piggy-backs onto NF-κB/RELA and utilizes the κB-motif at a cis-regulatory region to control mir-21 expression. p53 behaves as a co-factor in this complex because despite a mutation in its DNA binding domain, mutant p53 was still capable of binding RELA and the cis-element, and inducing mir-21 expression. In dilated human hearts where mir-21 upregulation was previously demonstrated, the p53-RELA complex was also associated with this cis-element. Using high-throughput sequencing, we analyzed genome-wide binding sites for the p53-RELA complex in diseased and control human hearts and found a significant overrepresentation of the STAT3 motif. We further determined that STAT3 was necessary for the p53-RELA complex to associate with this cis-element and for mir-21 expression.

Conclusions

Our results uncover a mechanism by which transcription factors cooperate in a multi-molecular complex at a cis-regulatory element to control gene expression.