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Nucleic Acids Research Advance Access published online on December 15, 2008
Nucleic Acids Research, doi:10.1093/nar/gkn978
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© 2008 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Gene regulation, Chromatin and Epigenetics

Identification of DNA regions and a set of transcriptional regulatory factors involved in transcriptional regulation of several human liver-enriched transcription factor genes

Hisashi Miura1,2, Yasuhiro Tomaru1,2, Misato Nakanishi1,2, Shinji Kondo3, Yoshihide Hayashizaki1,2,4 and Masanori Suzuki1,2,*

1RIKEN Omics Science Center (OSC), RIKEN Yokohama Institute 1-7-22 Suehiro-Cho, 2Division of Functional Genomics, Supramolecular Biology, International Graduate School of Arts and Sciences, Yokohama City University, 1-7-29 Suehiro-cho, Tsurumi-Ku, Yokohama, Kanagawa 230-0045, 3MetaSystems Research Team, Computational Systems Biology Research Group, Advanced Computational Sciences Department, RIKEN Advanced Science Institute and 4Genome Science Laboratory, Discovery and Research Institute, RIKEN Wako Main Campus, 2-1 Hirosawa, Wako, 351-0198, Japan

*To whom correspondence should be addressed. Tel: +81 045 508 7241; Fax: +81 045 508 7370; Email: msuzuki{at}gsc.riken.jp or msuzuki{at}tsurumi.yokohama-cu.ac.jp

Received August 18, 2008. Revised November 18, 2008. Accepted November 20, 2008.

Mammalian tissue- and/or time-specific transcription is primarily regulated in a combinatorial fashion through interactions between a specific set of transcriptional regulatory factors (TRFs) and their cognate cis-regulatory elements located in the regulatory regions. In exploring the DNA regions and TRFs involved in combinatorial transcriptional regulation, we noted that individual knockdown of a set of human liver-enriched TRFs such as HNF1A, HNF3A, HNF3B, HNF3G and HNF4A resulted in perturbation of the expression of several single TRF genes, such as HNF1A, HNF3G and CEBPA genes. We thus searched the potential binding sites for these five TRFs in the highly conserved genomic regions around these three TRF genes and found several putative combinatorial regulatory regions. Chromatin immunoprecipitation analysis revealed that almost all of the putative regulatory DNA regions were bound by the TRFs as well as two coactivators (CBP and p300). The strong transcription-enhancing activity of the putative combinatorial regulatory region located downstream of the CEBPA gene was confirmed. EMSA demonstrated specific bindings of these HNFs to the target DNA region. Finally, co-transfection reporter assays with various combinations of expression vectors for these HNF genes demonstrated the transcriptional activation of the CEBPA gene in a combinatorial manner by these TRFs.


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