The LIM domains of hic-5 protein recognize specific DNA fragments in a zinc-dependent manner in vitro.
hic-5 protein is a member of the LIM protein family, containing four LIM domains in its C-terminal region. It is mainly localized in focal adhesions and shows striking similarity to paxillin in its LIM domains, although the function of these LIM domains has remained elusive. In the present study, we found that full-length and the C-terminal half of hic-5 protein, including four LIM domains, bound to DNA in a zinc-dependent manner in vitro . Mouse genomic fragments that specifically bound to the hic-5 protein were isolated by successive rounds of hic-5 protein-DNA complex immunoprecipitation and PCR amplification. Seven independent clones were isolated, which contained high amounts of G+A and/or a long A/T tract. A DNA binding protein blot assay revealed the specificity of the interaction between hic-5 protein and the DNA fragment. Using a series of truncated forms of the hic-5 LIM domains, each of the four LIM domains was found to contribute to DNA binding in a distinctive manner.
Department of Microbiology, Showa University School of Pharmaceutical Sciences, Hatanodai 1-5-8, Shinagawa-ku, Tokyo, Japan and Osaka Bioscience Institute, Suita, Osaka, Japan.
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Nishiya
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Sabe
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Nose
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Shibanuma
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D003598:Cytoskeletal Proteins / Q000737:chemistry* / Q000378:metabolism*; D004247:DNA / Q000737:chemistry* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D004926:Escherichia coli; D007700:Kinetics; D060588:LIM Domain Proteins; D051379:Mice; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D015032:Zinc / Q000494:pharmacology*; D016335:Zinc Fingers
D003598:Cytoskeletal Proteins; D004268:DNA-Binding Proteins; D060588:LIM Domain Proteins; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; C090090:Tgfb1i1 protein, mouse; D004247:DNA; D015032:Zinc