The long-term objective of this proposal is to develop novel tools for studying how repair of double strand breaks by sister chromatid recombination is regulated in the context of mouse development. This work is made possible by two novel tools developed in cell culture models: first, a novel recombination reporter that allows quantification of specific sister chromatid recombination outcomes by flow cytometry;second, the development of a system for the tight regulation of the rare-cutting restriction endonuclease, I-SceI. This proposal aims to develop mouse strains carrying the recombination reporter expressed from the ubiquitously expressed ROSA26 locus and inducible alleles of I-SceI expressed from the ColA1 locus. These models will provide powerful new tools for studying the relationship between homologous recombination/sister chromatid recombination, aging and cancer.
Our specific aims are: 1. Generate a breeding colony of sister chromatid recombination reporter mice. 2. Generate a breeding colony of I-SceI-inducible mice. 3. Cross these strains to generate inducible recombination reporter mice. 4. Assess the performance of these reporter strains in sister chromatid recombination both ex vivo and in vivo.
Project Narrative The accurate repair of chromosome breaks by a process termed """"""""sister chromatid recombination"""""""" (SCR) is critical for preventing chromosome errors, and defects in this process contribute to aging and cancer. To assess how SCR is regulated in different tissues of the body, or during the development of a living mammal, we have developed and validated powerful new tools for studying sister chromatid recombination using flow cytometry. Here, we will use these same tools to begin to study how SCR is regulated in a live mouse.
| Chandramouly, Gurushankar; Kwok, Amy; Huang, Bin et al. (2013) BRCA1 and CtIP suppress long-tract gene conversion between sister chromatids. Nat Commun 4:2404 |
| Hartlerode, Andrea; Odate, Shobu; Shim, Inbo et al. (2011) Cell cycle-dependent induction of homologous recombination by a tightly regulated I-SceI fusion protein. PLoS One 6:e16501 |
| Scully, Ralph (2011) Epistatic relationships in the BRCA1-BRCA2 pathway. PLoS Genet 7:e1002183 |
| Scully, Ralph (2010) A histone code for DNA repair. Nat Rev Mol Cell Biol 11:164 |
| Scully, Ralph (2010) The spindle-assembly checkpoint, aneuploidy, and gastrointestinal cancer. N Engl J Med 363:2665-6 |
