1. Stalled replication forks within heterochromatin require ATRX for protection.
- Author
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Huh MS, Ivanochko D, Hashem LE, Curtin M, Delorme M, Goodall E, Yan K, and Picketts DJ
- Subjects
- Animals, Ataxia Telangiectasia Mutated Proteins genetics, Ataxia Telangiectasia Mutated Proteins metabolism, BRCA1 Protein, Carrier Proteins genetics, Cell Proliferation drug effects, Co-Repressor Proteins, DNA genetics, DNA metabolism, DNA Damage, DNA Helicases deficiency, DNA Repair Enzymes genetics, DNA Repair Enzymes metabolism, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, Gene Expression Regulation, HeLa Cells, Heterochromatin drug effects, Heterochromatin metabolism, Homeodomain Proteins genetics, Homeodomain Proteins metabolism, Humans, Hydroxyurea pharmacology, Intracellular Signaling Peptides and Proteins deficiency, Intracellular Signaling Peptides and Proteins genetics, MRE11 Homologue Protein, Mice, Mice, Knockout, Molecular Chaperones, Nerve Tissue Proteins genetics, Nerve Tissue Proteins metabolism, Neurogenesis genetics, Neurons cytology, Neurons drug effects, Nuclear Proteins deficiency, Nuclear Proteins metabolism, POU Domain Factors genetics, POU Domain Factors metabolism, Poly (ADP-Ribose) Polymerase-1 metabolism, Prosencephalon cytology, Prosencephalon drug effects, Prosencephalon metabolism, Rad51 Recombinase genetics, Rad51 Recombinase metabolism, Repressor Proteins genetics, Repressor Proteins metabolism, Tumor Suppressor Proteins genetics, Tumor Suppressor Proteins metabolism, X-linked Nuclear Protein, DNA Helicases genetics, DNA Replication, Heterochromatin chemistry, Neurons metabolism, Nuclear Proteins genetics, Poly (ADP-Ribose) Polymerase-1 genetics
- Abstract
Expansive growth of neural progenitor cells (NPCs) is a prerequisite to the temporal waves of neuronal differentiation that generate the six-layered neocortex, while also placing a heavy burden on proteins that regulate chromatin packaging and genome integrity. This problem is further reflected by the growing number of developmental disorders caused by mutations in chromatin regulators. ATRX gene mutations cause a severe intellectual disability disorder (α-thalassemia mental retardation X-linked (ATRX) syndrome; OMIM no. 301040), characterized by microcephaly, urogenital abnormalities and α-thalassemia. Although the ATRX protein is required for the maintenance of repetitive DNA within heterochromatin, how this translates to disease pathogenesis remain poorly understood and was a focus of this study. We demonstrate that Atrx(FoxG1Cre) forebrain-specific conditional knockout mice display poly(ADP-ribose) polymerase-1 (Parp-1) hyperactivation during neurogenesis and generate fewer late-born Cux1- and Brn2-positive neurons that accounts for the reduced cortical size. Moreover, DNA damage, induced Parp-1 and Atm activation is elevated in progenitor cells and contributes to their increased level of cell death. ATRX-null HeLa cells are similarly sensitive to hydroxyurea-induced replication stress, accumulate DNA damage and proliferate poorly. Impaired BRCA1-RAD51 colocalization and PARP-1 hyperactivation indicated that stalled replication forks are not efficiently protected. DNA fiber assays confirmed that MRE11 degradation of stalled replication forks was rampant in the absence of ATRX or DAXX. Indeed, fork degradation in ATRX-null cells could be attenuated by treatment with the MRE11 inhibitor mirin, or exacerbated by inhibiting PARP-1 activity. Taken together, these results suggest that ATRX is required to limit replication stress during cellular proliferation, whereas upregulation of PARP-1 activity functions as a compensatory mechanism to protect stalled forks, limiting genomic damage, and facilitating late-born neuron production.
- Published
- 2016
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