Exploration of cis-regulatory diversity underlying phenotypic innovation

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Project Summary The falling costs of DNA sequencing has resulted in a proliferation of genome-wide association studies (GWAS) seeking to identify causal variants associated with disease and other important phenotypes. However, a striking 93% of phenotype-associated variants fall within non-coding regions, often several hundreds of kilobases away from the nearest gene. Recent studies have demonstrated that such variants are enriched within cis-regulatory elements (CREs) and can affect transcriptional outcomes. CREs are composed of clusters of 4-30 bp DNA motifs recognized by sequence-specific transcription factors (TFs) that cooperatively establish patterns of transcription in a development and cell type-specific manner. However, it remains unclear how genetic variants within CREs mechanistically perturb patterns of transcription and contribute towards phenotypic diversity at the scale of individual cells, tissues and whole organisms. The aim of this proposal is to determine the molecular relationships among genetic variants, CREs, and gene transcription across diverse cell types and genetic backgrounds, and their concerted effects on cellular and organismal phenotypes. Zea mays (maize) is characterized by extensive intraspecies phenotypic and genetic variation comparable to the levels observed among primates. The genetic framework of this proposal, the 282 maize diversity panel, was specifically constructed from geographically dispersed individuals to represent the full spectrum of extant variation within the species. The rate of linkage-disequilibrium decay in the 282 maize diversity panel is 10-40X that of human and mouse genomes, affording significantly smaller population sizes with equivalent resolution. The rationale for this study is that the abundance of genetic variation, genomic resources, reduced population size requirements and recent expansion of reference-quality genomes (~35) present an ideal model to investigate the mechanistic basis of phenotypic diversity stemming from regulatory variation. The proposed research is innovative in that cutting edge genomic approaches, including scATAC-seq and snRNA-seq, will be utilized in parallel with microscopy-based imaging across 200 maize genotypes to test the central hypothesis that CRE variation underlies distinct spatiotemporal patterns of gene transcription that collectively manifest in phenotypic diversity at cellular and organismal levels. Realization of the proposed aims will usher a new understanding of the determinants of cell type-specificity and the phenotypic consequences of evolving gene regulatory landscapes. Successful completion the proposed work will lay the foundation for future interrogation of the mechanistic role of genetic variants towards human disease, relevant to the missions of the NIH. This application was specifically designed to enhance the applicants career development through associated training activities in mentorship, grant writing, and scientific communication. Accomplishment of the career training plan will facilitate the successful transition of the applicant from a post-doctoral position to a faculty position at a research-intensive institution.