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Molecular and Cellular Biology, March 2000, p. 2108-2121, Vol. 20, No. 6
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Cloning of a Mammalian Transcriptional Activator That Binds Unmethylated CpG Motifs and Shares a CXXC Domain with DNA Methyltransferase, Human Trithorax, and Methyl-CpG Binding Domain Protein 1

Kui Shin Voo, Diana L. Carlone, Britta M. Jacobsen, Anna Flodin, and David G. Skalnik*

Herman B. Wells Center for Pediatric Research and Section of Pediatric Hematology/Oncology, Departments of Pediatrics and Biochemistry & Molecular Biology, Indiana University School of Medicine, Indianapolis, Indiana 46202

Received 10 December 1999/Accepted 21 December 1999

Ligand screening was utilized to isolate a human cDNA that encodes a novel CpG binding protein, human CpG binding protein (hCGBP). This factor contains three cysteine-rich domains, two of which exhibit homology to the plant homeodomain finger domain. A third cysteine-rich domain conforms to the CXXC motif identified in DNA methyltransferase, human trithorax, and methyl-CpG binding domain protein 1. A fragment of hCGBP that contains the CXXC domain binds to an oligonucleotide probe containing a single CpG site, and this complex is disrupted by distinct oligonucleotide competitors that also contain a CpG motif(s). However, hCGBP fails to bind oligonucleotides in which the CpG motif is either mutated or methylated, and it does not bind to single-stranded DNA or RNA probes. Furthermore, the introduction of a CpG dinucleotide into an unrelated oligonucleotide sequence is sufficient to produce a binding site for hCGBP. Native hCGBP is detected as an 88-kDa protein by Western analysis and is ubiquitously expressed. The DNA-binding activity of native hCGBP is apparent in electrophoretic mobility shift assays, and hCGBP trans-activates promoters that contain CpG motifs but not promoters in which the CpG is ablated. These data indicate that hCGBP is a transcriptional activator that recognizes unmethylated CpG dinucleotides, suggesting a role in modulating the expression of genes located within CpG islands.

* Corresponding author. Mailing address: Wells Center for Pediatric Research, Cancer Research Building, Room 472, 1044 W. Walnut St., Indianapolis, IN 46202. Phone: (317) 274-8977. Fax: (317) 274-8928. E-mail: dskalnik{at}iupui.edu.

Molecular and Cellular Biology, March 2000, p. 2108-2121, Vol. 20, No. 6
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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