Loss of Atrx Sensitizes Cells to DNA Damaging Agents through p53-Mediated Death Pathways

Damiano Conte,Emma Goodall,Michael Huh,Marilyne Delorme,Robin J Parks,David J Picketts,Gilbert Bernier

doi:10.1371/journal.pone.0052167

Damiano Conte, Emma Goodall + Show 5 more

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https://doi.org/10.1371/journal.pone.0052167

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Journal: PloS one	Publication Date: Dec 17, 2012
Citations: 31	License type: CC BY 4.0

Affiliation: Ottawa Hospital, University of Ottawa

Abstract

Prevalent cell death in forebrain- and Sertoli cell-specific Atrx knockout mice suggest that Atrx is important for cell survival. However, conditional ablation in other tissues is not associated with increased death indicating that diverse cell types respond differently to the loss of this chromatin remodeling protein. Here, primary macrophages isolated from Atrx f/f mice were infected with adenovirus expressing Cre recombinase or β-galactosidase, and assayed for cell survival under different experimental conditions. Macrophages survive without Atrx but undergo rapid apoptosis upon lipopolysaccharide (LPS) activation suggesting that chromatin reorganization in response to external stimuli is compromised. Using this system we next tested the effect of different apoptotic stimuli on cell survival. We observed that survival of Atrx-null cells were similar to wild type cells in response to serum withdrawal, anti-Fas antibody, C2 ceramide or dexamethasone treatment but were more sensitive to 5-fluorouracil (5-FU). Cell survival could be rescued by re-introducing Atrx or by removal of p53 demonstrating the cell autonomous nature of the effect and its p53-dependence. Finally, we demonstrate that multiple primary cell types (myoblasts, embryonic fibroblasts and neurospheres) were sensitive to 5-FU, cisplatin, and UV light treatment. Together, our results suggest that cells lacking Atrx are more sensitive to DNA damaging agents and that this may result in enhanced death during development when cells are at their proliferative peak. Moreover, it identifies potential treatment options for cancers associated with ATRX mutations, including glioblastoma and pancreatic neuroendocrine tumors.

Highlights

Constitutional mutations in the ATRX gene cause a rare form of X-linked intellectual disability, namely the a-thalassemia mental retardation syndrome (ATR-X; OMIM# 30032) [1]
Studies in the forebrain have suggested that Atrx is critical for cell survival, acting through a p53-dependent pathway [20,22]
To overcome these difficulties and explore the mechanisms contributing to cell death in Atrx mutant mice we utilized cells harvested from Atrxf/f mice followed by infection with adenoviruses expressing either bgalactosidase (AdLacZ) as a control, or Cre-recombinase (AdCre) to inactivate the Atrx gene

Summary

Introduction

Constitutional mutations in the ATRX gene cause a rare form of X-linked intellectual disability, namely the a-thalassemia mental retardation syndrome (ATR-X; OMIM# 30032) [1]. Most disease causing mutations are missense changes located within two highly conserved regions, an N-terminal ADD domain (an atypical PHD domain common to ATRX, DNMT3 and DNMT3L) and a C-terminal ATPase/ helicase motif shared by the many Swi2/Snf2-like chromatin remodeling proteins. These two domains define the known biochemical properties and functions of the ATRX protein. Daxx-ATRX complexes are necessary for the deposition of the histone variant H3.3 at pericentromeric and telomeric heterochromatin [13,14,15] Despite these advances in our understanding of ATRX biochemical function it is not clear how these activities contribute to disease pathology

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