Biochemical Characterization of BRCA2
Abstract
The cell lines derived from BRCA2 mutant mice (complete knockout and C-terminus trancated mutant) are hypersensitive to the DNA crosslinking agent, mitomycin C (MMC). MMC induces DNA interstand crosslinks (crosslinks), which is repaired b an ill-defined crosslink repair pathway. Our hypothesis is that BRCA2 directly participates in the removal of crosslinks by forming a DNA repair complex with other factors. Our plan is to isolate a BRCA2-crosslink repair complex using our cell-free crosslink repair assay. In order to accomplish this goal, we need to prepare cell free extract from BRCA2 mutant cells. There are currently three sources of BRCA2 mutant cells, MEF cells from BRCA2 mutant mice, CAPAN-l cells and XRCC-ll Chinese hamster mutant cells. We chose to use CAPAN-l cell because CAPAN-l cells are available from ATCC. We examined the sensitivity of CAPAN-l cells to MMC and DV radiation. Similar to the reported phenotypes or mouse and hamster BRCA2 mutant cells, CAPAN-l cells are sensitive to MMC, but not to DV irradiation. The data showed that CAPAN-l cells are most likely defective in crosslink repair. We are currently growing CAPAN-l cells in a large scale to prepare cell-free extract. Once we prepare cell-free extract, we will proceed with the proposed experiments to isolate a BRCA2-containing crosslink repair complex.
Document Details
- Document Type
- Technical Report
- Publication Date
- Apr 01, 2004
- Accession Number
- ADA425128
Entities
People
- Tadayoshi Bessho
Organizations
- University of Texas Health Science Center at San Antonio