DNA-binding profiling of human hormone nuclear receptors via fluorescence correlation spectroscopy in a cell-free system

Kobayashi, Tamiyo; Kodani, Yoshiko; Nozawa, Akira; Endo, Yaeta; Sawasaki, Tatsuya

doi:10.1016/j.febslet.2008.07.003

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Volume 582, Issue 18 , Pages 2737-2744, 6 August 2008

DNA-binding profiling of human hormone nuclear receptors via fluorescence correlation spectroscopy in a cell-free system

Edited by Paul Bertone

Tamiyo Kobayashi
- Cell-Free Science and Technology Research Center, and The Venture Business Laboratory, Ehime University, 3 Bunkyo-cho, Matsuyama, Ehime 790-8577, Japan
- Life Science Group, Olympus Corp., 2-3 Kuboyama-cho, Hachioji, Tokyo 192-8512, Japan
Yoshiko Kodani
- Cell-Free Science and Technology Research Center, and The Venture Business Laboratory, Ehime University, 3 Bunkyo-cho, Matsuyama, Ehime 790-8577, Japan
Akira Nozawa
- Cell-Free Science and Technology Research Center, and The Venture Business Laboratory, Ehime University, 3 Bunkyo-cho, Matsuyama, Ehime 790-8577, Japan
- RIKEN Genomic Sciences Center, 1-7-22 Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan
Yaeta Endo
- Cell-Free Science and Technology Research Center, and The Venture Business Laboratory, Ehime University, 3 Bunkyo-cho, Matsuyama, Ehime 790-8577, Japan
- RIKEN Genomic Sciences Center, 1-7-22 Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan
- Corresponding authors. Address: Cell-Free Science and Technology Research Center, Venture Business Laboratory, Ehime University, 3 Bunkyo-cho, Matsuyama, Ehime 790-8577, Japan. Fax: +81 89 927 9941 (T. Sawasaki).
Tatsuya Sawasaki
- Cell-Free Science and Technology Research Center, and The Venture Business Laboratory, Ehime University, 3 Bunkyo-cho, Matsuyama, Ehime 790-8577, Japan
- RIKEN Genomic Sciences Center, 1-7-22 Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan
- Corresponding authors. Address: Cell-Free Science and Technology Research Center, Venture Business Laboratory, Ehime University, 3 Bunkyo-cho, Matsuyama, Ehime 790-8577, Japan. Fax: +81 89 927 9941 (T. Sawasaki).

Received 11 April 2008; received in revised form 1 July 2008; accepted 1 July 2008. published online 10 July 2008.

Abstract

The nuclear hormone receptors (NHRs), a family of transcription factors, bind directly to the hormone response elements (HREs) to regulate gene expression. In this study, we describe a comprehensive NHR–HRE profiling analysis with a new high-throughput DNA binding assay system utilizing wheat germ cell-free protein production and fluorescence correlation spectroscopy (FCS). This approach revealed NHR binding to natural response elements and new heterodimeric NHR–HRE bindings. We analyzed 408 possible binding combinations between 34 human NHRs and 12 different HREs, and identified 205 NHR–HRE binding combinations, 124 of which have not been previously reported. Thus, this study provides a novel biochemical classification of the human NHRs, as well as describing a novel approach to the large-scale analysis of DNA–protein interactions.

Keywords: Cell-free protein production, DNA-binding profiling, Fluorescence correlation spectroscopy, Nuclear hormone receptor, Hormone response elements, Biochemical annotation