Project Summary Clonal expansions increase with age throughout many tissues in the body. These outgrowths are driven by somatic mutations that provide a selective advantage over wild-type cells within a specific microenvironment. Clonal expansions radically increase with age, and are also shaped by disease-related factors. How these factors and clonal mosaicism (CM), reciprocally influences each other constitute some of the most important biological questions in current human genetics. A detailed understanding of the biology of aging at the cellular and molecular level is needed for the development of potential mitigation therapies that target aging morbidities. To make progress toward this goal, we need enhanced ability to identify and characterize recurrent somatic mutations in both healthy and diseased aging tissues at high-resolution. As identification of clones is hampered by low tissue availability and resource-intensive DNA sequencing methods, conceptual and technical innovation overcoming these challenges could rapidly accelerate the pace of variant discovery. Once discovered, functional characterization of these clonal variants in genetically heterogenous tissues requires application of genotype- aware, multi-modality methods in primary human samples to link genotypes and epigenetic profiles of individual cells. Currently, no such tools exist to characterize multiple molecular features of the somatic epigenome. To address these important gaps, we aim to (i) develop tools for the identification of candidate somatic variants at scale from publicly available data and (ii) apply novel technologies that map genotype to epigenetic phenotype in single cells, enabling analysis of multiple tissues from clinical samples obtained from aging individuals. Specifically, we aim to harness advancements in machine learning to develop a tool that can accurately identify somatic variants in bulk RNA-seq and apply it to publicly available data in healthy and diseased tissue from aged individuals. We believe that we can gain valuable information about the clonal architecture of diabetes, obesity, hypertension and other age-related diseases from our public databases at low cost and high speed. Furthermore, we aim to examine how epigenetic modifier mutations shape the chromatin regulatory landscape (including histone modifications and chromatin accessibility) to promote clonal outgrowth in CM. We will use newly developed tools to assay genotype, chromatin accessibility and histone marks in ASXL1-mutant CH and FOXO1-mutant fatty liver disease to define differentially accessible and active regions, their associated genes and transcription factor binding motifs and their enrichment along lineage trajectories. By integrating these modalities, we will have an unprecedented view of the effect of somatic variants on the aging epigenome and gain insights into how these mutations may drive clonal expansion. This project will be ideal for a training physician-scientist,...