Altered DNA binding specificity of Arnt by selection of partner bHLH-PAS proteins

Koshi Kinoshita, Yasuo Kikuchi, Yukie Sasakura, Masashi Suzuki, Yoshiaki Fujii-Kuriyama, Kazuhiro Sogawa

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    16 Citations (Scopus)

    Abstract

    The Ah receptor (AhR) and HLF are transcription factors involved in xenobiotic metabolism and hypoxic response, respectively. AhR and HLF heterodimerize with Arnt as the common partner, and bind to asymmetric E-boxes termed XRE and HRE, respectively. In order to investigate nucleotide preference of the heterodimers, reporter plasmids with oligonucleotides for XREs or HREs with systematic mutations were constructed and their activity was determined. Comparison of the activity revealed that DNA length and nucleotide preference recognized by Arnt subunit in the two heterodimers were largely different between XRE and HRE. We expressed AhR-Arnt and HLF-Arnt in Escherichia coli and used them for DNA binding. The dissociation constant of HLF-Arnt-HRE was 10.4 ± 1.6 nM. Competition activity of mutated XREs or HREs with wild type was consistent with their transcription activity. Bending of XRE and HRE induced by binding of the relevant heterodimers was observed with stronger bending of XRE than of HRE. By deletional and mutational analyses, an alanine and three arginine (Ala 8, Arg 9, Arg 11 and Arg 12) residues in the basic sequence of HLF were found to be indispensable for the transcriptional activity.

    Original languageEnglish
    Pages (from-to)3169-3179
    Number of pages11
    JournalNucleic acids research
    Volume32
    Issue number10
    DOIs
    Publication statusPublished - 2004 Aug 16

    ASJC Scopus subject areas

    • Genetics

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  • Cite this

    Kinoshita, K., Kikuchi, Y., Sasakura, Y., Suzuki, M., Fujii-Kuriyama, Y., & Sogawa, K. (2004). Altered DNA binding specificity of Arnt by selection of partner bHLH-PAS proteins. Nucleic acids research, 32(10), 3169-3179. https://doi.org/10.1093/nar/gkh637