The Human Enhancer Blocker CTC-binding Factor Interacts with the Transcription Factor Kaiso

Pierre-Antoine Defossez,Kevin F Kelly,Guillaume J.P Filion,Roberto Pérez-Torrado,Frédérique Magdinier,Hervé Menoni,Curtis L Nordgaard,Juliet M Daniel,Eric Gilson

doi:10.1074/jbc.m510802200

Abstract

CTC-binding factor (CTCF) is a DNA-binding protein of vertebrates that plays essential roles in regulating genome activity through its capacity to act as an enhancer blocker. We performed a yeast two-hybrid screen to identify protein partners of CTCF that could regulate its activity. Using full-length CTCF as bait we recovered Kaiso, a POZ-zinc finger transcription factor, as a specific binding partner. The interaction occurs through a C-terminal region of CTCF and the POZ domain of Kaiso. CTCF and Kaiso are co-expressed in many tissues, and CTCF was specifically co-immunoprecipitated by several Kaiso monoclonal antibodies from nuclear lysates. Kaiso is a bimodal transcription factor that recognizes methylated CpG dinucleotides or a conserved unmethylated sequence (TNGCAGGA, the Kaiso binding site). We identified one consensus unmethylated Kaiso binding site in close proximity to the CTCF binding site in the human 5' beta-globin insulator. We found, in an insulation assay, that the presence of this Kaiso binding site reduced the enhancer-blocking activity of CTCF. These data suggest that the Kaiso-CTCF interaction negatively regulates CTCF insulator activity.

Highlights

CTCF was originally isolated as a zinc-finger transcription factor that recognized a CTC-rich sequence in the c-myc promoter [6]
Kaiso is a member of the POZ family of zinc finger (ZF) transcription factors that are implicated in cancer and development [16]
Because POZ domains are highly conserved and are present in a large number of transcription factors [30], we tested the specificity of the interaction by asking whether CTCF would interact with the POZ domain of four other POZ-ZF proteins (BCL-6, PLZF, HIC-1, FAZF)

Summary

Introduction

CTCF was originally isolated as a zinc-finger transcription factor that recognized a CTC-rich sequence in the c-myc promoter [6]. An additional role of CTCF is to act as an enhancer blocker that prevents communication between an enhancer and a target gene This process is known as transcriptional insulation. Significant advances were made in our understanding of how CTCF functions as an enhancer blocker at the 5Ј chicken ␤-globin insulator [12, 13] At this insulator site, CTCF interacts with nucleophosmin, a nuclear matrix protein that is concentrated at the surface of the nucleolus. CTCF interacts with nucleophosmin, a nuclear matrix protein that is concentrated at the surface of the nucleolus This is thought to result in the formation of physically separated DNA loops, which would prevent an enhancer element in one chromatin loop from acting on a gene in the neighboring chromatin loop. This raises the possibility that Kaiso is a negative regulator of CTCF enhancer-blocking activity

Methods

Results

Conclusion