title: An evolutionarily conserved DNA architecture determines target specificity of the TWIST family bHLH transcription factors.
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.
authors.affiliation: Department of Pharmacology, University of California at San Diego, La Jolla, California, 92093, USA; The Biomedical Sciences Graduate Program, University of California at San Diego, La Jolla, California, 92093, USA;
authors.forename: Andrew T
authors.initials: AT
authors.lastname: Chang
authors.affiliation: The Wistar Institute, Philadelphia, Pennsylvania 19104, USA;
authors.forename: Yuanjie
authors.initials: Y
authors.lastname: Liu
authors.affiliation: The Wistar Institute, Philadelphia, Pennsylvania 19104, USA;
authors.forename: Kasirajan
authors.initials: K
authors.lastname: Ayyanathan
authors.affiliation: Salk Institute for Biological Studies, La Jolla, California 92037, USA;
authors.forename: Chris
authors.initials: C
authors.lastname: Benner
authors.affiliation: Department of Pharmacology, University of California at San Diego, La Jolla, California, 92093, USA; The Biological Science Graduate Program, University of California at San Diego, La Jolla, California, 92093, USA;
authors.forename: Yike
authors.initials: Y
authors.lastname: Jiang
authors.affiliation: Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA;
authors.forename: Jeremy W
authors.initials: JW
authors.lastname: Prokop
authors.affiliation: Department of Pharmacology, University of California at San Diego, La Jolla, California, 92093, USA;
authors.forename: Helicia
authors.initials: H
authors.lastname: Paz
authors.affiliation: Department of Cell and Molecular Medicine, University of California at San Diego, La Jolla, California, 92093, USA;
authors.forename: Dong
authors.initials: D
authors.lastname: Wang
authors.affiliation: Department of Cell and Molecular Medicine, University of California at San Diego, La Jolla, California, 92093, USA;
authors.forename: Hai-Ri
authors.initials: HR
authors.lastname: Li
authors.affiliation: Department of Cell and Molecular Medicine, University of California at San Diego, La Jolla, California, 92093, USA;
authors.forename: Xiang-Dong
authors.initials: XD
authors.lastname: Fu
authors.affiliation: The Wistar Institute, Philadelphia, Pennsylvania 19104, USA; jingyang@ucsd.edu rauscher@wistar.org.
authors.forename: Frank J
authors.initials: FJ
authors.lastname: Rauscher
authors.affiliation: Department of Pharmacology, University of California at San Diego, La Jolla, California, 92093, USA; Department of Pediatrics, University of California at San Diego, La Jolla, California, 92093, USA jingyang@ucsd.edu rauscher@wistar.org.
authors.forename: Jing
authors.initials: J
authors.lastname: Yang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D005075:Biological Evolution; D017124:Conserved Sequence; D004330:Drosophila / Q000737:chemistry / Q000378:metabolism; D029721:Drosophila Proteins / Q000737:chemistry* / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D008958:Models, Molecular; D011485:Protein Binding; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D051879:Twist-Related Protein 1 / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D029721:Drosophila Proteins; D051879:Twist-Related Protein 1
keywords: TWIST; WR domain; bHLH transcription factor; epithelial–mesenchymal transition