A Family of Human Zinc Finger Proteins That Bind Methylated DNA and Repress Transcription

Abstract

In vertebrates, densely methylated DNA is associated with inactive transcription. Actors in this process include proteins of the MBD family that can recognize methylated CpGs and repress transcription. Kaiso, a structurally unrelated protein, has also been shown to bind methylated CGCGs through its three Krüppel-like C₂H₂ zinc fingers. The human genome contains two uncharacterized proteins, ZBTB4 and ZBTB38, that contain Kaiso-like zinc fingers. We report that ZBTB4 and ZBTB38 bind methylated DNA in vitro and in vivo. Unlike Kaiso, they can bind single methylated CpGs. When transfected in mouse cells, the proteins colocalize with foci of heavily methylated satellite DNA and become delocalized upon loss of DNA methylation. Chromatin immunoprecipitation suggests that both of these proteins specifically bind to the methylated allele of the H19/Igf2 differentially methylated region. ZBTB4 and ZBTB38 repress the transcription of methylated templates in transfection assays. The two genes have distinct tissue-specific expression patterns, but both are highly expressed in the brain. Our results reveal the existence of a family of Kaiso-like proteins that bind methylated CpGs. Like proteins of the MBD family, they are able to repress transcription in a methyl-dependent manner, yet their tissue-specific expression pattern suggests nonoverlapping functions.

We are especially grateful to Philippe Ravassard and Jacques Mallet for assistance with the realization and interpretation of the in situ hybridization experiment, to Howard Cedar for the dnmt1 ^{− / −} cell line, to Brian Hendrich for the Mbd2^−/− cell line, and to Wolf Reik for the gift of Sd7 mice. We thank the following colleagues who contributed plasmids and other reagents: Alexander Bershadsky, Howard Cedar, Juliet Daniel, Phillip James, Francesca Lembo, Dominique Leprince, and Roger Tsien. We thank Dominique Leprince, Richard Meehan, and Fyodor Urnov for helpful comments on the manuscript.

S.S. and E.P. were supported by the Wellcome Trust, grant GR067436MA. S.Z. was supported by a grant from the Russian Foundation for Basic Research. Work in the Defossez lab was supported by the Curie Institute, CNRS (program ATIP), Fondation pour la Recherche Médicale, and Association pour la Recherche contre le Cancer.