DNA Nucleotide Excision Repair-Dependent Signaling to Checkpoint Activation

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 103; no. 46; pp. 17325 - 17330
• Main Authors: Marini, Federica, Nardo, Tiziana, Giannattasio, Michele, Minuzzo, Mario, Stefanini, Miria, Plevani, Paolo, Falconi, Marco Muzi
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 14-11-2006; National Acad Sciences
• Subjects: Antibodies; Biological Sciences; Cell cycle; Cell lines; Cells, Cultured; Deoxyribonucleic acid; DNA; DNA - genetics; DNA damage; DNA repair; DNA Repair - genetics; Eukaryotes; Fibroblasts; Genetic mutation; Genome, Human - genetics; Humans; Irradiation; Lesions; Mutation; Phosphorylation; Proteins; Radiation damage; Signal Transduction; Studies; Transcription, Genetic - genetics; Ultraviolet radiation; Xeroderma Pigmentosum - genetics
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0605446103

Eukaryotic cells respond to a variety of DNA insults by triggering a common signal transduction cascade, known as checkpoint response, which temporarily halts cell-cycle progression. Although the main players involved in the cascade have been identified, there is still uncertainty about the nature of the structures that activate these surveillance mechanisms. To understand the role of nucleotide excision repair (NER) in checkpoint activation, we analyzed the UV-induced phosphorylation of the key checkpoint proteins Chkl and p53, in primary fibroblasts from patients with xeroderma pigmentosum (XP), Cockayne syndrome (CS), trichothiodystrophy (TTD), or UV light-sensitive syndrome. These disorders are due to defects in transcription-coupled NER (TC-NER) and/or global genome NER (GG-NER), the NER subpathways repairing the transcribed strand of active genes or the rest of the genome, respectively. We show here that in G₀/G₁ and G₂/M phases of the cell cycle, triggering of the DNA damage cascade requires recognition and processing of the lesions by the GG-NER. Loss of TC-NER does not affect checkpoint activation. Mutations in XPD, XPB, and in TTDA, encoding subunits of the TFIIH complex, involved in both transcription and NER, impair checkpoint triggering. The only exception is represented by mutations in XPD, resulting in combined features of XP and CS (XP/CS) that lead to activation of the checkpoint cascade after UV radiation. Inhibition of RNA polymerase II transcription significantly reduces the phosphorylation of key checkpoint factors in XP/CS fibroblasts on exposure to UV damage.