Collaborative Research: The Epigenetic Regulatory Role of the Maize Ufo1 Gene

智力优势：允许形式和功能多样化的机制之一是表观遗传调节。表观遗传变化是影响基因表达的染色质的DNA或蛋白质组分的改变。具有瞬时性质的表观遗传修饰在植物界中广泛存在，但很少被分离或鉴定，因为相关的表型不容易看到，并且修饰在一代又一代之间不稳定。利用玉米果皮颜色1（p1）基因及其红色色素沉着作为标记，p1位点的等位基因变异已被证明是基因结构和组织的功能，这是进一步调节表观遗传机制。 为了理解通过基因表达的表观遗传调节来调节等位基因多样性的基础，已经进行了被称为Ufo 1（橙色不稳定因子1）的显性突变的分子和遗传表征。具有特定p1等位基因（称为P1-wr）的植物产生具有无色籽粒果皮和红色穗轴颖片的穗，并且这种沉默果皮表型是稳定的;然而，在Ufo 1存在下，P1-wr等位基因赋予在几种组织中色素沉着增强的表型。 表达研究表明，色素沉着的增益与P1转录的稳态水平增加有关。P1-wr; Ufo 1植物中P1表达增强与P1-wr等位基因启动子和编码序列DNA甲基化降低相关。 在本项目中，将进行ufo 1基因的图位克隆。利用转座子诱变将产生ufo 1的新等位基因。Ufo 1介导的表观遗传调控的影响的证据将使用高通量测序方法进行分析的染色质，转录本（mRNA和小RNA），和DNA甲基化的Ufo 1突变体和野生型植物的全球概况。将开发一个数据库，以可视化和分析ChIP-seq、RNA-seq和亚硫酸氢盐测序生成的数据。结果将确定Ufo 1突变（和野生型ufo 1基因）在影响转座子、DNA甲基化、组蛋白修饰和RNA谱的全局变化中的作用。由于Ufo 1在玉米中的作用方式与其他已知的表观遗传调节因子不同，因此该项目将扩大对重叠表观遗传途径如何调节这一重要作物基因表达的理解。更广泛的影响：该项目将提供植物表观遗传学和植物基因等位变异研究的培训机会。宾夕法尼亚州立大学和特拉华州大学之间的合作将加强植物生物学和生物信息学领域的专业互动。具体而言，两名博士后研究员和两名研究生将在计算生物学方面和表观遗传基因调控方面进行交叉培训。本科生，包括妇女和少数民族学生，将学习经典和尖端的植物生物学技术，用于理解和剖析组织特异性基因表达调控的分子基础。此外，高中学生和教师将参加暑期生物技术讲习班，学习玉米基因表达技术。该项目由分子和细胞生物科学部的遗传机制计划和综合有机系统部的植物基因组研究计划共同资助。

Intellectual Merit: One of the mechanisms that allow diversification of form and function is epigenetic regulation. Epigenetic changes are alterations in the DNA or protein components of chromatin that affect gene expression. Epigenetic modifications that are of transient nature are widespread in the plant kingdom, but are rarely isolated or identified because the associated phenotypes are not easy to see and the modifications are not stable from generation to generation. Using the pericarp color1 (p1) gene and its red pigmentation as a marker in maize, allelic variation at the p1 locus has been shown to be a function of gene structure and organization, which is further regulated by epigenetic mechanisms. To understand the basis of allelic diversity modulated through epigenetic regulation of gene expression, molecular and genetic characterization of a dominant mutation called Ufo1 (Unstable factor for orange1) has been performed. Plants with a particular p1 allele, called P1-wr, produce ears with colorless kernel pericarps and red cob glumes, and this silent pericarp phenotype is stable; however, in the presence of Ufo1 the P1-wr allele confers variably enhanced pigmentation phenotypes in several tissues. Expression studies demonstrated that the gain of pigmentation is associated with increased steady state levels of P1 transcript. Enhanced P1 expression in P1-wr;Ufo1 plants was shown to correlate with a decrease in DNA methylation at the promoter and coding sequence of the P1-wr allele. In the current project, map based cloning of ufo1 gene will be carried out. New alleles of ufo1 will be generated using transposon mutagenesis. Evidence for Ufo1-mediated effects on epigenetic regulation will be analyzed using high throughput sequencing methods for global profiling of chromatin, transcript (mRNA and small RNA), and DNA methylation of Ufo1 mutant and wild type plants. A database will be developed to visualize and analyze data generated from ChIP-seq, RNA-seq and bisulfite sequencing. The results will establish the role of the Ufo1 mutation (and wild type ufo1 gene) in effecting global changes in transposons, DNA methylation, histone modifications and RNA profiles. Because Ufo1 does not appear to act in the same way as other known epigenetic regulators in maize, this project will expand understanding of how overlapping epigenetic pathways regulate gene expression in this important crop plant.Broader Impacts: This project will provide training opportunities in plant epigenetics and study of allelic variation of plant genes. The collaboration between Pennsylvania State University and the University Delaware will enhance professional interactions in the area of plant biology and bioinformatics. Specifically, two postdoctoral fellows and two graduate students will be cross-trained in computational biological aspects and epigenetic gene regulation. Undergraduate students, including women and minority students, will learn classical and cutting-edge plant biology techniques that are used understand and dissect the molecular basis of regulation of tissue-specific gene expression. In addition, high school students and teachers will participate in a summer biotechnology workshop to learn gene expression techniques in maize.This project is co-funded by the Genetic Mechanisms Program in the Division of Molecular and Cellular Biosciences and by the Plant Genome Research Program in the Division of Integrative Organismal Systems.