S-phase sensing of DNA-protein crosslinks triggers TopBP1-independent ATR activation and p53-mediated cell death by formaldehyde

• Published in: Cell cycle (Georgetown, Tex.) Vol. 11; no. 13; pp. 2526 - 2537
• Main Authors: Wong, Victor Chun-Lam, Cash, Haley L., Morse, Jessica, Lu, Shan, Zhitkovich, Anatoly
• Format: Journal Article
• Language: English
• Published: United States Taylor & Francis 01-07-2012; Landes Bioscience
• Subjects: apoptosis; Apoptosis - drug effects; Ataxia Telangiectasia Mutated Proteins; ATR; Binding; Biology; Bioscience; Calcium; Cancer; Carrier Proteins - metabolism; Cell; Cell Cycle Proteins - metabolism; Cell Line; Checkpoint Kinase 1; Cycle; DNA - metabolism; DNA Adducts - chemistry; DNA Adducts - metabolism; DNA Damage - drug effects; DNA Mismatch Repair - drug effects; DNA-Binding Proteins - metabolism; DNA-protein crosslink; formaldehyde; Formaldehyde - pharmacology; HCT116 Cells; Humans; Landes; Nuclear Proteins - metabolism; Organogenesis; Phosphorylation; Protein Kinases - metabolism; Protein-Serine-Threonine Kinases - metabolism; Proteins; RNA Interference; RNA, Small Interfering - metabolism; S Phase; senescence; Signal Transduction; Tumor Suppressor Protein p53 - antagonists & inhibitors; Tumor Suppressor Protein p53 - genetics; Tumor Suppressor Protein p53 - metabolism
• ISSN: 1538-4101; 1551-4005
• DOI: 10.4161/cc.20905

We examined genotoxic signaling and cell fate decisions in response to a potent DNA-protein crosslinker formaldehyde (FA). DNA-protein crosslinks (DPC) are poorly understood lesions produced by bifunctional carcinogens and several cancer drugs. FA-treated human cells showed a rapid activation of ATR kinase that preferentially targeted the p53 transcription factor at low doses and CHK1 kinase at more severe damage, producing bell-shaped and sublinear responses, respectively. CHK1 phosphorylation was transient, and its loss was accompanied by increased p53 accumulation and Ser15 phosphorylation. Activation of p53 was insensitive to inhibition of mismatch repair and nucleotide and base excision repair, excluding the role of small DNA adducts in this response. The p53-targeted signaling was transcription-independent, absent in quiescent cells and specific to S-phase in cycling populations. Unlike other S-phase stressors, FA-activated p53 was functional transcriptionally, promoted apoptosis in lung epithelial cells and caused senescence in normal lung fibroblasts. FA did not induce ATR, RAD1 or RPA foci, and p53 phosphorylation was TopBP1-independent, indicating a noncanonical mode of ATR activation. Replication arrest by FA caused a dissociation of ATR from a chromatin-loaded MCM helicase but no PCNA monoubiquitination associated with stalled polymerases. These results suggest that unlike typical DNA adducts that stall DNA polymerases, replication inhibition by bulkier DPC largely results from blocking upstream MCM helicase, which prevents accumulation of ssDNA. Overall, our findings indicate that S-phase-specific, TopBP1-independent activation of the ATR-p53 axis is a critical stress response to FA-DPC, which has implications for understanding of FA carcinogenesis.