Burden and signatures of somatic mutations in genomes of healthy individuals.

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Somatic mutations accumulate over the lifetime of an individual due to both genetic and environmental factors. It is becoming evident that somatic genome changes are associated with a host of pathologies including cancers. Sequencing genomes of different cancer types suggested that mutation loads vary between cell types and across the body. The variations have been associated with differential exposure to DNA damaging agents and the replicative potential of the cells. In addition, mutation loads due to DNA damaging lesions would also be dependent on the ability of the cells to repair damage in an error-free manner. However, the mutation loads attributable to environmental and intrinsic factors across cell types in healthy individuals are not known. Also, it is not known how polymorphisms within DNA repair genes compromise repair efficiency and alter the mutation landscape in cells exposed to environmental DNA damage as well as in unexposed cells. The goal of this proposal is to determine the extent of somatic genome changes within the body and in different individuals and to examine the mechanisms that contribute to this variability. To address this goal, I will explore the following aims. In Aim1, I will directly analyze the impact of DNA repair polymorphisms associated with cancers on DNA repair capacity using orthogonal systems. Using plasmid-based host cell reactivation assays, I will test repair efficiency in lymphocytes with homozygous minor (mutant) or major (wild-type) alleles. I will also determine if these mutant human genes increase mutation and recombination rates in yeast and human cells upon exposure to exogenous DNA damage and during unchallenged growth. In Aim2, I will determine the role of deleterious single nucleotide polymorphisms (SNPs) in DNA repair genes. Mutations leading to loss of a functional MBD4 glycosylase, have been shown to increase CT changes in CpG dinucleotides in cancer genomes. I will test if SNPs that are predicted to be deleterious to the MBD4 protein also increase mutation loads and altering signatures in somatic cells from healthy individuals. In Aim3, I will estimate mutation loads in different cell-types isolated from the same individuals from different body sites. I will assess the contributions of mutation signatures associated with known environmental and endogenous mutagenic sources to mutation loads in the samples. The completion of the studies in this proposal will provide me with expertise in cell culture, genetic manipulation of human cell lines and bioinformatics, paving the way for a successful career as an independent researcher. Significance: These studies will increase our understanding of the interplay of environmental and genetic factors that determine somatic mutagenesis. These results are important for understanding the susceptibility of individuals to cancers and other diseases associated with somatic mutagenesis, and in designing individual- specific disease prevention strategies.