Aging and DNA RepairCurrent hypothesis of aging is that it results from accumulation of damage to macromolecules. Among various macromolecules DNA is the only one that cannot be recycled if damaged. To deal with DNA damage organisms have evolved elaborate DNA repair machinery. The repair, however, is not perfect and often results in mutations. Mutations in DNA accumulate over time leading to cancer and aging. In addition to simple point mutations old cells accumulate specific type of mutations: large genomic rearrangements. These rearrangements result from errors made by DNA double-stranded break (DSB) repair machinery. Therefore, we believe that DNA DSB repair plays an important role in the aging process. DNA DSBs are repaired by two pathways: homologous recombination (HR) and by Nonhomologous End Joining (NHEJ). HR typically copies the missing information from the sister chromatid into the break site, resulting in exact reconstitution of the original sequence. In contrast, NHEJ fuses the two broken ends, with little regard for homology, leading to deletions and other rearrangements. We have shown that DNA repair by NHEJ becomes less efficient and more error-prone in senescent human cells. We are now investigating the molecular mechanisms responsible for the decline of NHEJ during aging. We showed that the level of Ku protein, which plays a key role in NHEJ declines in senescent human cells. We are also studying age-related changes in the second pathway of DSB repair, homologous recombination. We also work on developing transgenic mouse model for analysis of DNA double-strand break repair in an aging organism. Our goal is to characterize the changes that occur in DSB repair during aging and cellular senescence, identify the mechanisms responsible for these changes, and to find ways to improve the function of DNA repair in an aging organism.
We developed sensitive reporter assays, which we use to measure the efficiency of NHEJ and HR. The reporter cassettes are based on green fluorescent protein (GFP). DNA DSBs can be induced in specific location inside the GFP gene and repair results in reactivation of GFP activity (green cells).
Normal human fibroblasts co-transfected with NHEJ reporter cassette and pDsRed.
Content last modified: May 13, 2013