Exploration of cis-regulatory diversity underlying phenotypic innovation

项目摘要 DNA测序成本的下降导致了全基因组关联研究（GWAS）的激增 寻求确定与疾病和其他重要表型相关的致病变异。然而，一个引人注目的 93%的表型相关变异属于非编码区，通常距离数百个碱基 最接近的基因。最近的研究表明，这些变异体在顺式调节基因中富集， 元件（克雷斯），并可以影响转录结果。克雷斯由4-30 bp的DNA基序簇组成 由序列特异性转录因子（TF）识别，这些转录因子合作建立转录模式 以发育和细胞类型特异性的方式。然而，目前尚不清楚克雷斯中的遗传变异如何影响基因表达。 机械地扰乱转录模式，并有助于表型多样性的规模 单个细胞、组织和整个生物体。这项建议的目的是确定分子关系 在不同细胞类型和遗传背景中的遗传变异、克雷斯和基因转录，以及 它们对细胞和有机体表型的协同作用。 玉米具有广泛的种内表型和遗传变异， 达到灵长类动物的水平这项建议的遗传框架，282玉米多样性小组， 是专门从地理上分散的个人，以代表现有的全谱 物种内的变异。在282个玉米多样性群体中连锁不平衡衰减率为10- 40倍 人类和小鼠的基因组，提供了相当的分辨率明显较小的人口规模。 这项研究的理由是，丰富的遗传变异，基因组资源，减少人口 参考质量基因组（~35）的大小要求和最近的扩展提供了一个理想的模型， 研究由调节变异引起的表型多样性的机制基础。拟议 研究是创新的，因为尖端的基因组方法，包括scATAC-seq和snRNA-seq，将被 在200个玉米基因型中与基于显微镜的成像并行使用，以检验中心假设 CRE变异是基因转录的不同时空模式的基础， 细胞和生物体水平的表型多样性。实现拟议的目标将迎来一个新的 理解细胞类型特异性的决定因素和进化基因的表型后果 监管景观。顺利完成了所提出的工作，为以后的审讯工作打下了基础 遗传变异对人类疾病的机械作用，与NIH的使命有关。 此应用程序是专门设计的，以提高申请人的职业发展，通过相关的 在指导、赠款写作和科学交流方面的培训活动。职业成就 培训计划将有助于申请人从博士后职位成功过渡到教师 在一家研究密集型机构工作。

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.