DNA Damage Changes Distribution Pattern and Levels of HP1 Protein Isoforms in the Nucleolus and Increases Phosphorylation of HP1β-Ser88.

Legartová Legartová,Pokorná Pokorná,Kozubek Kozubek,Lochmanová Lochmanová,Šponer Šponer,Krepl Krepl,Zdráhal Zdráhal,Bártová Bártová

doi:10.3390/cells8091097

Abstract

The family of heterochromatin protein 1 (HP1) isoforms is essential for chromatin packaging, regulation of gene expression, and repair of damaged DNA. Here we document that γ-radiation reduced the number of HP1α-positive foci, but not HP1β and HP1γ foci, located in the vicinity of the fibrillarin-positive region of the nucleolus. The additional analysis confirmed that γ-radiation has the ability to significantly decrease the level of HP1α in rDNA promoter and rDNA encoding 28S rRNA. By mass spectrometry, we showed that treatment by γ-rays enhanced the HP1β serine 88 phosphorylation (S88ph), but other analyzed modifications of HP1β, including S161ph/Y163ph, S171ph, and S174ph, were not changed in cells exposed to γ-rays or treated by the HDAC inhibitor (HDACi). Interestingly, a combination of HDACi and γ-radiation increased the level of HP1α and HP1γ. The level of HP1β remained identical before and after the HDACi/γ-rays treatment, but HDACi strengthened HP1β interaction with the KRAB-associated protein 1 (KAP1) protein. Conversely, HP1γ did not interact with KAP1, although approximately 40% of HP1γ foci co-localized with accumulated KAP1. Especially HP1γ foci at the periphery of nucleoli were mostly absent of KAP1. Together, DNA damage changed the morphology, levels, and interaction properties of HP1 isoforms. Also, γ-irradiation-induced hyperphosphorylation of the HP1β protein; thus, HP1β-S88ph could be considered as an important marker of DNA damage.

Highlights

The chromatin of the eukaryotic cell nucleus is a dynamic and highly organized complex consisting of DNA and histones
We addressed a question if irradiation by γ-rays changes the morphology of heterochromatin protein 1 (HP1)-positive foci and their localization close to nucleoli (Figure 1a–d)
Due to the results showing an increased HP1β-serine 88 phosphorylation (S88ph) in γ-irradiated HeLa cells, and these cells treated by SAHA (Figure 4a), we addressed a question if HDAC inhibitor, SAHA, induces DNA damage

Summary

Introduction

The chromatin of the eukaryotic cell nucleus is a dynamic and highly organized complex consisting of DNA and histones. In the interphase cell nucleus, chromatin appears as two primary and distinct isoforms referred to as euchromatin and heterochromatin. Heterochromatin is gene-poor, highly condensed, and mostly remains transcriptionally silent throughout the cell cycle, replicates in the late S phase, which is the attribute of repetitive sequences. Constitutive heterochromatin preferentially consists of repetitive elements such as satellite DNA in centromeres or telomeric sequences [1,2,3]. In regions of constitutive heterochromatin, histones are mostly hypo-acetylated and, for example, hypermethylated on histone H3, lysine 9 (H3K9me2/me). In regions of constitutive heterochromatin, histones are mostly hypo-acetylated and, for example, hypermethylated on histone H3, lysine 9 (H3K9me2/me3)

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