title: Regulation of the activation of the Fanconi anemia pathway by the p21 cyclin-dependent kinase inhibitor.
abstract: Fanconi anemia (FA) is a rare disease characterized by congenital defects, progressive bone marrow failure and heightened cancer susceptibility. The FA proteins, BRCA1 and FANCD1/BRCA2 function cooperatively in the FA-BRCA pathway to repair damaged DNA. Activation of the FA-BRCA pathway occurs via the monoubiquitination of the FANCD2 and FANCI proteins, targeting these proteins to discrete nuclear foci where they function in DNA repair. The cellular regulation of FANCD2/I monoubiquitination, however, remains poorly understood. In this study, we have examined the roles of the p53 tumor suppressor protein, as well as its downstream target, the p21(Cip1/Waf1) cyclin-dependent kinase inhibitor, in the regulation of the activation of the FA-BRCA pathway. We demonstrate that, in contrast to p53, p21 has a major role in the regulation of the activation of the FA-BRCA pathway: p21 promotes S-phase and DNA damage-inducible FANCD2/I monoubiquitination and nuclear foci formation. Several lines of evidence establish that this effect is not a consequence of a defective G1-S checkpoint or altered cell-cycle progression in the absence of p21. Instead, we demonstrate that p21 is required for the transcriptional repression of the USP1 deubiquitinating enzyme upon exposure to DNA-damaging agents. In the absence of p21, persistent USP1 expression precludes the DNA damage-inducible accumulation of monoubiquitinated FANCD2 and FANCI. Consequently, p21(-/-) cells exhibit increased levels of mitomycin C-inducible complex chromosomal aberrations and elevated γH2AX nuclear foci formation. Our results demonstrate that p21 has a critical role in the regulation of the activation of the FA-BRCA pathway and suggest a broader role for p21 in the orchestration of DNA repair processes following exposure to DNA crosslinking agents.
authors.affiliation: Department of Cell and Molecular Biology, University of Rhode Island, Kingston, RI 02881, USA.
authors.forename: M A
authors.initials: MA
authors.lastname: Rego
authors.forename: J A
authors.initials: JA
authors.lastname: Harney
authors.forename: M
authors.initials: M
authors.lastname: Mauro
authors.forename: M
authors.initials: M
authors.lastname: Shen
authors.forename: N G
authors.initials: NG
authors.lastname: Howlett
mesh_terms: D019313:BRCA1 Protein / Q000378:metabolism; D024682:BRCA2 Protein / Q000378:metabolism; D002453:Cell Cycle; D002460:Cell Line; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism*; D004249:DNA Damage; D005199:Fanconi Anemia / Q000378:metabolism*; D052236:Fanconi Anemia Complementation Group D2 Protein / Q000378:metabolism; D051856:Fanconi Anemia Complementation Group Proteins / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D006801:Humans; D016685:Mitomycin / Q000009:adverse effects; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D054875:Ubiquitination
chemical_list: D019313:BRCA1 Protein; C492913:BRCA1 protein, human; D024682:BRCA2 Protein; C551750:BRCA2 protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; C499905:FANCD2 protein, human; C519921:FANCI protein, human; D052236:Fanconi Anemia Complementation Group D2 Protein; D051856:Fanconi Anemia Complementation Group Proteins; C468783:H2AX protein, human; D006657:Histones; D016159:Tumor Suppressor Protein p53; D016685:Mitomycin