title: Activation of DNA damage repair pathways in response to nitrogen mustard-induced DNA damage and toxicity in skin keratinocytes.
abstract: Nitrogen mustard (NM), a structural analog of chemical warfare agent sulfur mustard (SM), forms adducts and crosslinks with DNA, RNA and proteins. Here we studied the mechanism of NM-induced skin toxicity in response to double strand breaks (DSBs) resulting in cell cycle arrest to facilitate DNA repair, as a model for developing countermeasures against vesicant-induced skin injuries. NM exposure of mouse epidermal JB6 cells decreased cell growth and caused S-phase arrest. Consistent with these biological outcomes, NM exposure also increased comet tail extent moment and the levels of DNA DSB repair molecules phospho H2A.X Ser139 and p53 Ser15 indicating NM-induced DNA DSBs. Since DNA DSB repair occurs via non homologous end joining pathway (NHEJ) or homologous recombination repair (HRR) pathways, next we studied these two pathways and noted their activation as defined by an increase in phospho- and total DNA-PK levels, and the formation of Rad51 foci, respectively. To further analyze the role of these pathways in the cellular response to NM-induced cytotoxicity, NHEJ and HRR were inhibited by DNA-PK inhibitor NU7026 and Rad51 inhibitor BO2, respectively. Inhibition of NHEJ did not sensitize cells to NM-induced decrease in cell growth and cell cycle arrest. However, inhibition of the HRR pathway caused a significant increase in cell death, and prolonged G2M arrest following NM exposure. Together, our findings, indicating that HRR is the key pathway involved in the repair of NM-induced DNA DSBs, could be useful in developing new therapeutic strategies against vesicant-induced skin injury.
authors.affiliation: Department of Pharmaceutical Sciences, University of Colorado Anchutz Medical Campus, Skaggs School of Pharmacy and Pharmaceutical Sciences, Aurora, CO 80045, USA.
authors.forename: Swetha
authors.initials: S
authors.lastname: Inturi
authors.affiliation: Department of Pharmaceutical Sciences, University of Colorado Anchutz Medical Campus, Skaggs School of Pharmacy and Pharmaceutical Sciences, Aurora, CO 80045, USA.
authors.forename: Neera
authors.initials: N
authors.lastname: Tewari-Singh
authors.affiliation: Department of Pharmaceutical Sciences, University of Colorado Anchutz Medical Campus, Skaggs School of Pharmacy and Pharmaceutical Sciences, Aurora, CO 80045, USA.
authors.forename: Chapla
authors.initials: C
authors.lastname: Agarwal
authors.affiliation: Department of Pediatrics, University of Colorado Anschutz Medical Campus, Aurora, CO 80045, USA.
authors.forename: Carl W
authors.initials: CW
authors.lastname: White
authors.affiliation: Department of Pharmaceutical Sciences, University of Colorado Anchutz Medical Campus, Skaggs School of Pharmacy and Pharmaceutical Sciences, Aurora, CO 80045, USA. Electronic address: Rajesh.Agarwal@ucdenver.edu.
authors.forename: Rajesh
authors.initials: R
authors.lastname: Agarwal
mesh_terms: D000818:Animals; D001896:Boron Compounds / Q000494:pharmacology; D002460:Cell Line; D002619:Chemical Warfare Agents / Q000633:toxicity*; D002867:Chromones / Q000494:pharmacology; D004249:DNA Damage; D004260:DNA Repair / Q000187:drug effects* / Q000235:genetics; D051747:DNA-Activated Protein Kinase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D059565:G2 Phase Cell Cycle Checkpoints / Q000187:drug effects / Q000235:genetics; D006657:Histones / Q000235:genetics / Q000378:metabolism; D015603:Keratinocytes / Q000378:metabolism* / Q000473:pathology; D008466:Mechlorethamine / Q000633:toxicity*; D051379:Mice; D009025:Morpholines / Q000494:pharmacology; D009687:Nuclear Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D051135:Rad51 Recombinase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D012867:Skin / Q000378:metabolism* / Q000473:pathology; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
chemical_list: D001896:Boron Compounds; D002619:Chemical Warfare Agents; D002867:Chromones; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins; D006657:Histones; D008466:Mechlorethamine; D009025:Morpholines; D009687:Nuclear Proteins; D051135:Rad51 Recombinase; D016159:Tumor Suppressor Protein p53; C479235:2-(morpholin-4-yl)benzo(h)chromen-4-one; C042168:boron oxide; C497264:Prkdc protein, mouse; C495154:Rad51 protein, mouse; C475961:gamma-H2AX protein, mouse
keywords: BO2; DNA double strand break (DSB) repair; HRR; NHEJ; NU7026; Nitrogen mustard; Rad51 inhibitor
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