Characterizing the Interplay between Maize Retrotransposons and the Epigenome

着丝粒是DNA片段，确保在分裂过程中将染色体忠实地分离到子细胞。着丝粒的结构还不是很清楚，尽管DNA和相关因子的协调变化是着丝粒功能所必需的。此外，着丝粒的结构和功能可能会随着时间的推移而改变，这引发了关于基因组如何在染色体水平上做出反应和适应的问题。着丝粒很难研究，因为它们由反转录转座子和其他移动元件组成，这些元件由高度重复的序列组成。组装短重复序列跨度的能力是出了名的困难，因此对研究人员来说，染色体的着丝粒区域仍然几乎是模糊的。该项目将使用新的测序技术和软件工具对所有10条玉米染色体的着丝粒区域进行测序和分析，这些技术和软件工具可以组装和分析重复序列，并可以研究着丝粒特有的相关因素。该项目的结果将增加已经测序的玉米基因组的价值，并将导致改进的参考基因组完全可供植物研究界使用。通过发现着丝粒的结构和功能，将扩大构建人工染色体的潜力，并可能识别与着丝粒相关的农艺性状。作为该项目的一部分，本科生将接受现代植物工程方法和合成生物学方面的培训。本科生团队将参加国际学生研究竞赛，从而促进学生和公众对植物着丝粒和合成生物学的了解。人们对DNA在着丝粒功能和进化中的作用知之甚少。在第一步中，将使用PacBio技术确定近交系B73参考基因组中所有10个玉米着丝粒的着丝粒特定重复的性质和排列。这些重复序列对着丝粒特异蛋白沉积的影响将使用许多不同的实验方法来确定，包括比较基因组学和表观基因组学，以及生化和遗传分析。新着丝粒的形成是一种表观遗传学变化，涉及着丝粒特异性组蛋白H3的CENH3移动到新的基因组区域。着丝粒反转录转座子(CR)对新着丝粒的快速入侵为研究表观遗传和遗传变化的相互作用提供了前所未有的机会。大量的表观遗传标记已经或正在被定位在标准玉米基因组中。组蛋白H2A.Z核小体可能代表转录区域的持久标记，将被绘制并与所有可用的表观遗传特征相结合，以确定遗传(双链DNA断裂和反转录转座子插入)和表观遗传(组蛋白变体、组蛋白修饰、基因表达、DNA甲基化)的相互影响。至少还有一个特殊结合着丝粒重复序列的表观遗传标记将被详细描述。在这项研究过程中产生的所有数据将保存在MaizeGDB(maizegdb.org)和GenBank(http://www.ncbi.nlm.nih.gov/genbank/).

Centromeres are DNA segments that ensure the faithful segregation of chromosomes to daughter cells during division. The structure of centromeres is not well understood, although coordinated changes in the DNA and associated factors are necessary for centromere function. Furthermore, centromere structure and function can change over time raising questions about how the genome responds and adapt at the chromosome level. Centromeres are difficult to study because they are composed of retrotranspons and other mobile elements that consist of highly repetitive sequences. The ability to assemble spans of short repeated sequences is notoriously difficult, so the centromere regions of chromosomes have remained nearly obscure to researchers. This project will sequence and analyze the centromere regions of all ten maize chromosomes using new sequencing technologies and software tools that permit assembly and analysis of the repetitive sequences and will permit study of the associated factors unique to centromeres. The results of this project will add value to the already sequenced maize genome and will result in an improved reference genome fully available to the plant research community. By discovering the structure and function of centromeres, the potential to construct artificial chromosomes will be expanded and agronomic traits associated with centromeres may be identified. As part of the project, undergraduate students will be trained in modern plant engineering methods and synthetic biology. The team of undergraduate students will participate in an international student research competition, thus promoting student and public understanding of plant centromeres and synthetic biology. Little is known about the role of DNA in centromere function and evolution. In a first step, the nature and arrangement of centromere-specific repeats at all ten maize centromeres of the inbred B73 reference genome will be determined using PacBio technology. The impact of these repeats on the deposition of centromere-specific proteins will be determined using a number of different experimental approaches, including comparative genomics and epigenomics, as well as biochemical and genetic assays. Neocentromere formation is an epigenetic change that involves movement of CENH3, the centromere specific histone H3, to a new genomic region. Rapid invasion of neocentromeres by centromeric retrotransposons (CR) provides an unprecedented opportunity to study the interplay of epigenetic and genetic changes. A large number of epigenetic marks have been, or are being, mapped in the standard maize genomes. Histone H2A.Z nucleosomes, which may represent persistent marks of transcribed regions, will be mapped and integrated with all available epigenetic signatures to determine the reciprocal effects of genetic (double-stranded DNA breaks and retrotransposons insertion) and epigenetic (histone variants, histone modifications, gene expression, DNA methylation). At least one additional epigenetic marker that specifically binds centromeric repeats will be characterized in detail. All data produced in the course of this research will be deposited at MaizeGDB (maizegdb.org) and GenBank (http://www.ncbi.nlm.nih.gov/genbank/).