title: ATM deficiency results in accumulation of DNA-topoisomerase I covalent intermediates in neural cells.
abstract: Accumulation of peptide-linked DNA breaks contributes to neurodegeration in humans. This is typified by defects in tyrosyl DNA phosphodiesterase 1 (TDP1) and human hereditary ataxia. TDP1 primarily operates at single-strand breaks (SSBs) created by oxidative stress or by collision of transcription machinery with topoisomerase I intermediates (Top1-CCs). Cellular and cell-free studies have shown that Top1 at stalled Top1-CCs is first degraded to a small peptide resulting in Top1-SSBs, which are the primary substrates for TDP1. Here we established an assay to directly compare Top1-SSBs and Top1-CCs. We subsequently employed this assay to reveal an increased steady state level of Top1-CCs in neural cells lacking Atm; the protein mutated in ataxia telangiectasia. Our data suggest that the accumulation of endogenous Top1-CCs in Atm-/- neural cells is primarily due to elevated levels of reactive oxygen species. Biochemical purification of Top1-CCs from neural cell extract and the use of Top1 poisons further confirmed a role for Atm during the formation/resolution of Top1-CCs. Finally, we report that global transcription is reduced in Atm-/- neural cells and fails to recover to normal levels following Top1-mediated DNA damage. Together, these data identify a distinct role for ATM during the formation/resolution of neural Top1-CCs and suggest that their accumulation contributes to the neuropathology of ataxia telangiectasia.
authors.affiliation: Genome Damage and Stability Centre, School of Life Sciences, University of Sussex, Brighton, United Kingdom.
authors.forename: Meryem
authors.initials: M
authors.lastname: Alagoz
authors.forename: Shih-Chieh
authors.initials: SC
authors.lastname: Chiang
authors.forename: Abhishek
authors.initials: A
authors.lastname: Sharma
authors.forename: Sherif F
authors.initials: SF
authors.lastname: El-Khamisy
mesh_terms: D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D064007:Ataxia Telangiectasia Mutated Proteins / Q000037:antagonists & inhibitors / Q000172:deficiency / Q000235:genetics*; D001681:Biological Assay; D002166:Camptothecin / Q000494:pharmacology; D002460:Cell Line; D053904:DNA Breaks, Single-Stranded; D004264:DNA Topoisomerases, Type I / Q000235:genetics* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D005786:Gene Expression Regulation; D006801:Humans; D009025:Morpholines / Q000494:pharmacology; D018384:Oxidative Stress; D010727:Phosphoric Diester Hydrolases / Q000235:genetics* / Q000378:metabolism; D011753:Pyrones / Q000494:pharmacology; D015398:Signal Transduction; D059004:Topoisomerase I Inhibitors / Q000494:pharmacology; D014158:Transcription, Genetic
chemical_list: C495818:2-morpholin-4-yl-6-thianthren-1-yl-pyran-4-one; D004791:Enzyme Inhibitors; D009025:Morpholines; D011753:Pyrones; D059004:Topoisomerase I Inhibitors; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D010727:Phosphoric Diester Hydrolases; C498508:TDP1 protein, human; D004264:DNA Topoisomerases, Type I; C528909:TOP1 protein, human; D002166:Camptothecin