James E.
Cleaver,
Ph.D.
University of California - San Francisco, UCSF Cancer Center
Endogenous DNA Damage and Mechanisms of Aging
Endogenous DNA damage and spontaneous mutagenesis may be important mechanisms in aging. We are developing mouse strains defective in base excision repair (BER) that cannot repair endogenous DNA damage through knockout of the Xrccl scaffold protein. The Xrccl protein interacts with ligase III to coordinate BER and is essential for viability in mice, but in Drosophila this interaction domain is missing and the flies are viable. We are constructing a vector that will allow us to terminate the mouse chromosomal gene immediately prior to the ligase III domain. This should allow other functions of the gene to continue and support a viable outcome. The BER-defective mouse strain would then be sensitive to developing degenerative changes associated with endogenous oxidative damage thought to be involved in aging. A second approach involves using the pro-mutagenic gene hRad30A that encodes a low-fidelity polymerase involved in the human disease xeroderma pigmentosum (XP). We have found that over-expression of hRad30A proves toxic to cells in culture, most likely by increasing spontaneous mutation rates. hRad30A can therefore be used to study the role of spontaneous mutagenesis in vivo in relation to aging, to complement studies in the BER knockout mouse. We are carrying out an initial investigation by expressing hRad30A on a keratin-14 promoter to increase its expression in the skin. If enhanced expression is pathological in the skin, we will next express the gene on a brain-specific promoter to develop a mouse model of neurodegeneration. The ultimate goal of these experiments is to develop mouse strains that show increased signs of aging due to spontaneous damage and mutagenesis, and use them for designing intervention strategies for improved human health.
