An evolutionarily conserved DNA architecture determines target specificity of the TWIST family bHLH transcription factors

  1. Jing Yang1,8
  1. 1Department of Pharmacology, University of California at San Diego, La Jolla, California, 92093, USA;
  2. 2The Biomedical Sciences Graduate Program, University of California at San Diego, La Jolla, California, 92093, USA;
  3. 3The Wistar Institute, Philadelphia, Pennsylvania 19104, USA;
  4. 4Salk Institute for Biological Studies, La Jolla, California 92037, USA;
  5. 5The Biological Science Graduate Program, University of California at San Diego, La Jolla, California, 92093, USA;
  6. 6Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA;
  7. 7Department of Cell and Molecular Medicine, University of California at San Diego, La Jolla, California, 92093, USA;
  8. 8Department of Pediatrics, University of California at San Diego, La Jolla, California, 92093, USA
  1. Corresponding authors: jingyang{at}ucsd.edu, rauscher{at}wistar.org
  1. 9 These authors contributed equally to this work.

Abstract

Basic helix–loop–helix (bHLH) transcription factors recognize the canonical E-box (CANNTG) to regulate gene transcription; however, given the prevalence of E-boxes in a genome, it has been puzzling how individual bHLH proteins selectively recognize E-box sequences on their targets. TWIST is a bHLH transcription factor that promotes epithelial–mesenchymal transition (EMT) during development and tumor metastasis. High-resolution mapping of TWIST occupancy in human and Drosophila genomes reveals that TWIST, but not other bHLH proteins, recognizes a unique double E-box motif with two E-boxes spaced preferentially by 5 nucleotides. Using molecular modeling and binding kinetic analyses, we found that the strict spatial configuration in the double E-box motif aligns two TWIST–E47 dimers on the same face of DNA, thus providing a high-affinity site for a highly stable intramolecular tetramer. Biochemical analyses showed that the WR domain of TWIST dimerizes to mediate tetramer formation, which is functionally required for TWIST-induced EMT. These results uncover a novel mechanism for a bHLH transcription factor to recognize a unique spatial configuration of E-boxes to achieve target specificity. The WR–WR domain interaction uncovered here sets an example of target gene specificity of a bHLH protein being controlled allosterically by a domain outside of the bHLH region.

Keywords

Footnotes

  • Received April 1, 2014.
  • Accepted February 9, 2015.

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