DNA damage response curtails detrimental replication stress and chromosomal instability induced by the dietary carcinogen PhIP.
PhIP is an abundant heterocyclic aromatic amine (HCA) and important dietary carcinogen. Following metabolic activation, PhIP causes bulky DNA lesions at the C8-position of guanine. Although C8-PhIP-dG adducts are mutagenic, their interference with the DNA replication machinery and the elicited DNA damage response (DDR) have not yet been studied. Here, we analyzed PhIP-triggered replicative stress and elucidated the role of the apical DDR kinases ATR, ATM and DNA-PKcsin the cellular defense response. First, we demonstrate that PhIP induced C8-PhIP-dG adducts and DNA strand breaks. This stimulated ATR-CHK1 signaling, phosphorylation of histone 2AX and the formation of RPA foci. In proliferating cells, PhIP treatment increased the frequency of stalled replication forks and reduced fork speed. Inhibition of ATR in the presence of PhIP-induced DNA damage strongly promoted the formation of DNA double-strand breaks, activation of the ATM-CHK2 pathway and hyperphosphorylation of RPA. The abrogation of ATR signaling potentiated the cell death response and enhanced chromosomal aberrations after PhIP treatment, while ATM and DNA-PK inhibition had only marginal effects. These results strongly support the notion that ATR plays a key role in the defense against cancer formation induced by PhIP and related HCAs.
Department of Toxicology, University Medical Center, Mainz, Germany.
Maximilian
M
Mimmler
Department of Toxicology, University Medical Center, Mainz, Germany.
Simon
S
Peter
Department of Toxicology, University Medical Center, Mainz, Germany.
Alexander
A
Kraus
Department of Toxicology, University Medical Center, Mainz, Germany.
Svenja
S
Stroh
Department of Toxicology, University Medical Center, Mainz, Germany.
Teodora
T
Nikolova
Department of Toxicology, University Medical Center, Mainz, Germany.
Nina
N
Seiwert
Department of Toxicology, University Medical Center, Mainz, Germany.
Solveig
S
Hasselwander
Department of Toxicology, University Medical Center, Mainz, Germany.
Carina
C
Neitzel
Department of Toxicology, University Medical Center, Mainz, Germany.
Jessica
J
Haub
Department of Food Safety, Federal Institute for Risk Assessment (BfR), Berlin, Germany.|Research Group Genotoxic Food Contaminants, German Institute of Human Nutrition (DIfE), Potsdam-Rehbrücke, Germany.
Bernhard H
BH
Monien
Institute for Food Toxicology and Analytical Chemistry, University of Veterinary Medicine Hannover, Hannover, Germany.
Petra
P
Nicken
Institute for Food Toxicology and Analytical Chemistry, University of Veterinary Medicine Hannover, Hannover, Germany.
Pablo
P
Steinberg
Department of Cell Biology, University of Texas Southwestern Medical Center, Dallas, TX, USA.
Jerry W
JW
Shay
Department of Toxicology, University Medical Center, Mainz, Germany.
Bernd
B
Kaina
Department of Toxicology, University Medical Center, Mainz, Germany fahrer@uni-mainz.de.
Jörg
J
Fahrer
D000818:Animals; D064007:Ataxia Telangiectasia Mutated Proteins / Q000378:metabolism; D002273:Carcinogens / Q000633:toxicity*; D002417:Cattle; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D000071877:Checkpoint Kinase 1 / Q000378:metabolism; D043171:Chromosomal Instability / Q000187:drug effects*; D002869:Chromosome Aberrations; D006224:Cricetinae; D018736:DNA Adducts; D053903:DNA Breaks, Double-Stranded; D004249:DNA Damage / Q000187:drug effects*; D004261:DNA Replication / Q000187:drug effects*; D000070818:Discoidin Domain Receptors / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007093:Imidazoles / Q000633:toxicity*; D010766:Phosphorylation; D015398:Signal Transduction / Q000187:drug effects; D013312:Stress, Physiological / Q000187:drug effects* / Q000235:genetics*
D002273:Carcinogens; D018736:DNA Adducts; D007093:Imidazoles; C049584:2-amino-1-methyl-6-phenylimidazo(4,5-b)pyridine; D000070818:Discoidin Domain Receptors; D064007:Ataxia Telangiectasia Mutated Proteins; D000071877:Checkpoint Kinase 1