Upon UV radiation or ionizing radiation, DNA replication forks can be stalled by inter-strand and intra-strand cross links. Stalled replication forks trigger assembly of the cellular DNA damage response machinery, which requires that some enzyme components of the DNA replication complex switch their physical locations, interaction partners and functions. One such component is FEN1 nuclease. Our preliminary data indicate that upon UV radiation, FEN1 switches from being a flap endonuclease for RNA primer removal and interacting with PCNA, to being a gap-dependent endonuclease for resolution of stalled DNA replication forks and interacting with WRN. This critical switch is mediated by a change in FEN1's post-translational modification (PTM) profile. These PTMs can act as a 'molecular barcode'that directs different FEN1-mediated protein- protein interactions to allow a switch of FEN1's functions. Disruption of the normal program of these PTMs may lead to uncontrolled cell growth and cancer. In this competitive renewal application, our goal is to establish a comprehensive relationship among genetic alterations, functional deficiency, and pathological consequences, using FEN1 nuclease as a model protein and transgenic mice as a model system. During the previous funding cycle we identified FEN1 mutations in cancer cells that eliminate the structural elements responsible for FEN1's nuclease activities, protein/protein interactions and PTMs. We also established corresponding knock-in mouse models that mimic the point mutations identified in human cancer, which enabled us to initially define the molecular and cellular events of tumorigenesis. These mouse models and our initial studies have prompted new aims to investigate the molecular mechanisms of FEN1mutation-mediated cancer pathogenesis caused by elevated mutagenesis during Okazaki fragment maturation and aberrant PTMs.
The Specific Aims i nclude: 1) To determine how FEN1-mediated 5'editing of ?-segment eliminates mispairs in Okazaki fragments and contributes to cancer avoidance. 2) To determine the role of FEN1/WRN complex in maintaining the stability of tandem repeat sequences and cancer avoidance. 3) To determine how UV radiation-induced post-translational modifications mediate a switch of FEN1's role from RNA primer removal to resolution of stalled replication forks. Successful completion of the proposed studies will generate important new knowledge about the function and regulation of FEN1 in maintenance of genome stabilities and cancer avoidance. In addition, the results are anticipated to have a high potential impact, as they may suggest new avenues for cancer prevention and development of new, personalized radiation and other therapeutic regimens for this life-threatening disease.
The current application aims to establish a comprehensive relationship among genetic alterations, functional deficiency, and pathological consequences, using human FEN1 nuclease as a model protein and transgenic mice as a model system. Successful completion of the proposed studies will gain important new knowledge about the function and regulation of flap endonuclease-1 in maintenance of genome stabilities and cancer avoidance. The results will serve as a reference in cancer prevention and development of new therapeutic regimens and personalized medicine.
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|Wang, Hailong; Li, Yongjiang; Truong, Lan N et al. (2014) CtIP maintains stability at common fragile sites and inverted repeats by end resection-independent endonuclease activity. Mol Cell 54:1012-21|
|Wu, Zhenxing; Lin, Yuanji; Xu, Hong et al. (2012) High risk of benzo[ýý]pyrene-induced lung cancer in E160D FEN1 mutant mice. Mutat Res 731:85-91|
|Zheng, Li; Dai, Huifang; Hegde, Muralidhar L et al. (2011) Fen1 mutations that specifically disrupt its interaction with PCNA cause aneuploidy-associated cancer. Cell Res 21:1052-67|
|Zheng, Li; Shen, Binghui (2011) Okazaki fragment maturation: nucleases take centre stage. J Mol Cell Biol 3:23-30|
|Tsutakawa, Susan E; Classen, Scott; Chapados, Brian R et al. (2011) Human flap endonuclease structures, DNA double-base flipping, and a unified understanding of the FEN1 superfamily. Cell 145:198-211|
|Xu, H; Zheng, L; Dai, H et al. (2011) Chemical-induced cancer incidence and underlying mechanisms in Fen1 mutant mice. Oncogene 30:1072-81|
|Zheng, Li; Jia, Jia; Finger, L David et al. (2011) Functional regulation of FEN1 nuclease and its link to cancer. Nucleic Acids Res 39:781-94|
|Guo, Zhigang; Zheng, Li; Xu, Hong et al. (2010) Methylation of FEN1 suppresses nearby phosphorylation and facilitates PCNA binding. Nat Chem Biol 6:766-73|
|Finger, L David; Blanchard, M Suzette; Theimer, Carla A et al. (2009) The 3'-flap pocket of human flap endonuclease 1 is critical for substrate binding and catalysis. J Biol Chem 284:22184-94|
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