Interaction of the epigenetic integrator UHRF1 with the MYST domain of TIP60 inside the cell

Waseem Ashraf,Tanveer Ahmad,Marc Mousli,Ludovic Richert,Mahmoud Alhosin,Ali Hamiche,Yves Mely,Liliyana Zaayter,Abdulkhaleg Ibrahim,Christian Bronner

doi:10.1186/s13046-017-0659-1

Abstract

BackgroundThe nuclear epigenetic integrator UHRF1 is known to play a key role with DNMT1 in maintaining the DNA methylation patterns during cell division. Among UHRF1 partners, TIP60 takes part in epigenetic regulations through its acetyltransferase activity. Both proteins are involved in multiple cellular functions such as chromatin remodeling, DNA damage repair and regulation of stability and activity of other proteins. The aim of this work was to investigate the interaction between UHRF1 and TIP60 in order to elucidate the dialogue between these two proteins.MethodsBiochemical (immunoprecipitation and pull-down assays) and microscopic (confocal and fluorescence lifetime imaging microscopy; FLIM) techniques were used to analyze the interaction between TIP60 and UHRF1 in vitro and in vivo. Global methylation levels were assessed by using a specific kit. The results were statistically analyzed using Graphpad prism and Origin.ResultsOur study shows that UHRF1, TIP60 and DNMT1 were found in the same epigenetic macro-molecular complex. In vitro pull-down assay showed that deletion of either the zinc finger in MYST domain or deletion of whole MYST domain from TIP60 significantly reduced its interaction with UHRF1. Confocal and FLIM microscopy showed that UHRF1 co-localized with TIP60 in the nucleus and confirmed that both proteins interacted together through the MYST domain of TIP60. Moreover, overexpression of TIP60 reduced the DNA methylation levels in HeLa cells along with downregulation of UHRF1 and DNMT1.ConclusionOur data demonstrate for the first time that TIP60 through its MYST domain directly interacts with UHRF1 which might be of high interest for the development of novel oncogenic inhibitors targeting this interaction.

Highlights

The nuclear epigenetic integrator UHRF1 is known to play a key role with DNA methyl transferase 1 (DNMT1) in maintaining the DNA methylation patterns during cell division
Immunoprecipitating UHRF1-mCherry by using anti-mCherry antibody led to the co-immunoprecipitation of both endogenous Tat-interacting protein 60 kDa (TIP60) and exogenous TIP60-Enhanced Green Fluorescent Protein (eGFP) while free eGFP which was co-transfected with UHRF1mCherry did not co-immunoprecipitate with it (Fig. 1c)
It is interesting to note that UHRF1-mCherry co-expression resulted in lower levels of TIP60-eGFP recombinant protein (Fig. 1d) as compared with cells transfected with TIP60-eGFP or co-transfected with mCherry alone

Summary

Introduction

The nuclear epigenetic integrator UHRF1 is known to play a key role with DNMT1 in maintaining the DNA methylation patterns during cell division. Among UHRF1 partners, TIP60 takes part in epigenetic regulations through its acetyltransferase activity. Both proteins are involved in multiple cellular functions such as chromatin remodeling, DNA damage repair and regulation of stability and activity of other proteins. Ubiquitin-like containing PHD and RING Finger domains 1 (UHRF1) is a multi-domain nuclear protein that plays an important role in epigenetics through the maintenance of DNA methylation patterns during DNA replication [1, 2]. UHRF1 is mostly up-regulated and its levels are maintained constant throughout the cell cycle. The high levels of UHRF1 found in variety of cancers are often

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