Uniform Widespread Nuclear Phosphorylation of Histone H2AX Is an Indicator of Lethal DNA Replication Stress
Phosphorylated histone H2AX (γ-H2AX), a central player in the DNA damage response (DDR), serves as a biomarker of DNA double-strand break repair. Although DNA damage is generally visualized by the formation of γ-H2AX foci in injured nuclei, it is unclear whether the widespread un...
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doaj-d104cea5417a4f12949f12648c9219f12020-11-25T02:17:23ZengMDPI AGCancers2072-66942019-03-0111335510.3390/cancers11030355cancers11030355Uniform Widespread Nuclear Phosphorylation of Histone H2AX Is an Indicator of Lethal DNA Replication StressEric Moeglin0Dominique Desplancq1Sascha Conic2Mustapha Oulad-Abdelghani3Audrey Stoessel4Manuela Chiper5Marc Vigneron6Pascal Didier7Laszlo Tora8Etienne Weiss9Biotechnologie et Signalisation Cellulaire, UMR 7242, CNRS/Université de Strasbourg, Boulevard S. Brant, 67412 Illlkirch, FranceBiotechnologie et Signalisation Cellulaire, UMR 7242, CNRS/Université de Strasbourg, Boulevard S. Brant, 67412 Illlkirch, FranceInstitut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch, FranceInstitut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch, FranceBiotechnologie et Signalisation Cellulaire, UMR 7242, CNRS/Université de Strasbourg, Boulevard S. Brant, 67412 Illlkirch, FranceBiotechnologie et Signalisation Cellulaire, UMR 7242, CNRS/Université de Strasbourg, Boulevard S. Brant, 67412 Illlkirch, FranceBiotechnologie et Signalisation Cellulaire, UMR 7242, CNRS/Université de Strasbourg, Boulevard S. Brant, 67412 Illlkirch, FranceLaboratoire de Bioimagerie et Pathologies, UMR 7213, CNRS/Université de Strasbourg, Route du Rhin, 67401 Illkirch, FranceInstitut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch, FranceBiotechnologie et Signalisation Cellulaire, UMR 7242, CNRS/Université de Strasbourg, Boulevard S. Brant, 67412 Illlkirch, FrancePhosphorylated histone H2AX (γ-H2AX), a central player in the DNA damage response (DDR), serves as a biomarker of DNA double-strand break repair. Although DNA damage is generally visualized by the formation of γ-H2AX foci in injured nuclei, it is unclear whether the widespread uniform nuclear γ-H2AX (called pan-nuclear) pattern occurring upon intense replication stress (RS) is linked to DDR. Using a novel monoclonal antibody that binds exclusively to the phosphorylated C-terminus of H2AX, we demonstrate that H2AX phosphorylation is systematically pan-nuclear in cancer cells stressed with RS-inducing drugs just before they die. The pan-nuclear γ-H2AX pattern is abolished by inhibition of the DNA-PK kinase. Cell death induction of cancer cells treated with increasing combinations of replication and kinase (ATR and Chk1) inhibitory drugs was proportional to the appearance of pan-nuclear γ-H2AX pattern. Delivery of labeled anti-γ-H2AX Fabs in stressed cells demonstrated at a single cell level that pan-nuclear γ-H2AX formation precedes irreversible cell death. Moreover, we show that H2AX is not required for RS-induced cell death in HeLa cells. Thus, the nuclear-wide formation of γ-H2AX is an incident of RS-induced cell death and, thus, the pan nuclear H2AX pattern should be regarded as an indicator of lethal RS-inducing drug efficacy.http://www.mdpi.com/2072-6694/11/3/355histone variantH2AX phosphorylationγ-H2AXpan-nuclear patternmonoclonal antibodycancer cellscell deathreplication stresschemotherapyH2AFX geneknock-out |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Eric Moeglin Dominique Desplancq Sascha Conic Mustapha Oulad-Abdelghani Audrey Stoessel Manuela Chiper Marc Vigneron Pascal Didier Laszlo Tora Etienne Weiss |
spellingShingle |
Eric Moeglin Dominique Desplancq Sascha Conic Mustapha Oulad-Abdelghani Audrey Stoessel Manuela Chiper Marc Vigneron Pascal Didier Laszlo Tora Etienne Weiss Uniform Widespread Nuclear Phosphorylation of Histone H2AX Is an Indicator of Lethal DNA Replication Stress Cancers histone variant H2AX phosphorylation γ-H2AX pan-nuclear pattern monoclonal antibody cancer cells cell death replication stress chemotherapy H2AFX gene knock-out |
author_facet |
Eric Moeglin Dominique Desplancq Sascha Conic Mustapha Oulad-Abdelghani Audrey Stoessel Manuela Chiper Marc Vigneron Pascal Didier Laszlo Tora Etienne Weiss |
author_sort |
Eric Moeglin |
title |
Uniform Widespread Nuclear Phosphorylation of Histone H2AX Is an Indicator of Lethal DNA Replication Stress |
title_short |
Uniform Widespread Nuclear Phosphorylation of Histone H2AX Is an Indicator of Lethal DNA Replication Stress |
title_full |
Uniform Widespread Nuclear Phosphorylation of Histone H2AX Is an Indicator of Lethal DNA Replication Stress |
title_fullStr |
Uniform Widespread Nuclear Phosphorylation of Histone H2AX Is an Indicator of Lethal DNA Replication Stress |
title_full_unstemmed |
Uniform Widespread Nuclear Phosphorylation of Histone H2AX Is an Indicator of Lethal DNA Replication Stress |
title_sort |
uniform widespread nuclear phosphorylation of histone h2ax is an indicator of lethal dna replication stress |
publisher |
MDPI AG |
series |
Cancers |
issn |
2072-6694 |
publishDate |
2019-03-01 |
description |
Phosphorylated histone H2AX (γ-H2AX), a central player in the DNA damage response (DDR), serves as a biomarker of DNA double-strand break repair. Although DNA damage is generally visualized by the formation of γ-H2AX foci in injured nuclei, it is unclear whether the widespread uniform nuclear γ-H2AX (called pan-nuclear) pattern occurring upon intense replication stress (RS) is linked to DDR. Using a novel monoclonal antibody that binds exclusively to the phosphorylated C-terminus of H2AX, we demonstrate that H2AX phosphorylation is systematically pan-nuclear in cancer cells stressed with RS-inducing drugs just before they die. The pan-nuclear γ-H2AX pattern is abolished by inhibition of the DNA-PK kinase. Cell death induction of cancer cells treated with increasing combinations of replication and kinase (ATR and Chk1) inhibitory drugs was proportional to the appearance of pan-nuclear γ-H2AX pattern. Delivery of labeled anti-γ-H2AX Fabs in stressed cells demonstrated at a single cell level that pan-nuclear γ-H2AX formation precedes irreversible cell death. Moreover, we show that H2AX is not required for RS-induced cell death in HeLa cells. Thus, the nuclear-wide formation of γ-H2AX is an incident of RS-induced cell death and, thus, the pan nuclear H2AX pattern should be regarded as an indicator of lethal RS-inducing drug efficacy. |
topic |
histone variant H2AX phosphorylation γ-H2AX pan-nuclear pattern monoclonal antibody cancer cells cell death replication stress chemotherapy H2AFX gene knock-out |
url |
http://www.mdpi.com/2072-6694/11/3/355 |
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