A local ATR-dependent checkpoint pathway is activated by a site-specific replication fork block in human cells Journal Article
| Authors: | Ahmed-Seghir, S.; Jalan, M.; Grimsley, H. E.; Sharma, A.; Twayana, S.; Kosiyatrakul, S. T.; Thompson, C.; Schildkraut, C. L.; Powell, S. N. |
| Article Title: | A local ATR-dependent checkpoint pathway is activated by a site-specific replication fork block in human cells |
| Abstract: | When replication forks encounter DNA lesions that cause polymerase stalling, a checkpoint pathway is activated. The ATR-dependent intra-S checkpoint pathway mediates detection and processing of sites of replication fork stalling to maintain genomic integrity. Several factors involved in the global checkpoint pathway have been identified, but the response to a single replication fork barrier (RFB) is poorly understood. We utilized the Escherichia coli-based Tus-Ter system in human MCF7 cells and showed that the Tus protein binding to TerB sequences creates an efficient site-specific RFB. The single fork RFB was sufficient to activate a local, but not global, ATR-dependent checkpoint response that leads to phosphorylation and accumulation of DNA damage sensor protein γH2AX, confined locally to within a kilobase of the site of stalling. These data support a model of local management of fork stalling, which allows global replication at sites other than the RFB to continue to progress without delay. © Ahmed-Seghir, Jalan et al. |
| Keywords: | signal transduction; controlled study; protein expression; protein phosphorylation; human cell; genetics; hydroxyurea; nonhuman; flow cytometry; dna replication; dna damage; cell cycle progression; gene overexpression; protein binding; immunofluorescence; phosphorylation; genetic transfection; chromatin immunoprecipitation; escherichia coli; western blotting; immunoprecipitation; plasmid; lentivirus vector; immunoblotting; reactive oxygen metabolite; atm protein; mitogen activated protein kinase 1; genomics; real time polymerase chain reaction; doxycycline; protein p21; atr protein; histone h2ax; atr protein, human; genomic dna; rna binding; pulsed field gel electrophoresis; gene replication; nucleosome; southern blotting; puromycin; phleomycin; humans; human; article; whole genome sequencing; ataxia telangiectasia mutated proteins; proteinase k; mcf-7 cell line; sodium hydroxide; crispr associated endonuclease cas9; benzylsulfonyl fluoride; cloxuridine |
| Journal Title: | eLife |
| Volume: | 12 |
| ISSN: | 2050-084X |
| Publisher: | eLife Sciences Publications Ltd. |
| Date Published: | 2023-08-30 |
| Start Page: | RP87357 |
| Language: | English |
| DOI: | 10.7554/eLife.87357 |
| PUBMED: | 37647215 |
| PROVIDER: | scopus |
| PMCID: | PMC10468204 |
| DOI/URL: | |
| Notes: | Article -- MSK Cancer Center Support Grant (P30 CA008748) acknowledged in PubMed and PDF -- MSK corresponding author is Simon Powell -- MSK author Sena Ahmed-Seghir's first name is misspelled on the publication -- Source: Scopus |
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