Tissue, Time, and Genotype: Investigating the Effects of Cellular Environment, Age, and Germline Mutation Rate on the Genome-Wide Rate and Spectrum of Somatic Mutation - Project Abstract
Somatic mutations represent a significant health concern as they underlie many cancers and
age-related disorders. Although technological advances have led to a deeper understanding of
germline mutations, somatic mutations have eluded deeper investigation until more recently.
The mutational process in germline versus somatic cells is likely similar, given both depend on
many of the same parameters (e.g., rates of DNA polymerase errors occurring during
replication and levels of DNA damage). In large multicellular organisms, however, somatic cells
differentiate during development and accumulate mutations due to repeated rounds of cell
division, thereby shifting these parameters across tissue types and over time. We propose to
investigate the relative influence of germline mutation rates, location, and age on somatic
mutation rates by performing whole genome sequencing on tissue-specific populations of cells
from multiple time points across genotypes for which a wide range of germline mutation has
already been observed. To our knowledge, this would be the first in organism comparison of
somatic and germline mutations across genotypes within a species. Our research strategy for
identifying somatic mutations will be PCR-free whole-genome DNA sequencing of cells sampled
from brain and heart, from juveniles and adults, across 9 genotypes of Daphnia magna, an
aquatic microcrustacean already in use for ageing research. Samples will be taken by
sectioning animals and using laser capture microdissection to isolate populations of cells from
specific tissues. High-throughput sequencing of multiplexed samples will allow us to efficiently
obtain sufficient (10x) coverage to call somatic variants across the genome. Continued
development of the D. magna system for investigating mutational processes allows us to (1)
leverage our current knowledge of germline mutation rates and phenotypic declines in this
species and (2) capitalize on our ability to quickly and easily target different tissue types in a
transparent arthropod, a long-standing model system in biology and public health research and
an emerging model system in genomics. Investigating the factors influencing variation in
somatic mutation rates will fill a major gap in our current knowledge of how DNA errors and
damage accumulates, providing insight into the frequency of mutation in different genomic
backgrounds, across tissues, and over time.
