Uncovering the molecular mechanism of paramutation, a classic example of non-Mendelian inheritance in maize

这个项目研究了一种被称为参数化的生物机制，它负责以高度可预测的方式打开和关闭选定的基因，并将这些“打开”和“关闭”状态传递给后代。与众所周知的突变过程类似，参数化产生影响进化成功的可遗传生物变异。然而，与突变不同的是，参数化的发生并不需要改变DNA密码。因此，该项目通过询问如何在没有DNA变化的情况下改变调控信息并将其传递给后代，解决了现代遗传学既定规则的一个例外。这项工作将在玉米上进行，玉米是研究参数化机制的卓越模型，因为负责植物颜色的基因的“开”和“关”状态可以直观地跟踪，控制交配很容易执行，并且可以生成和评估详细的多代谱系。在研究的基础上提出了一个参数化的工作模型，本项目对该模型的具体方面进行了测试。该项目还将确定在参数化过程中负责“开”和“关”开关的新分子和染色体结构。由于植物和动物都发生变异，这项研究的结果可能具有广泛的生物学影响，从增加对生殖生物学和遗传学的了解到开发改善农业或动物健康的新策略。将制作诸如彩色玉米穗之类的教育材料，并通过现有的推广计划在课堂上教授基本和高级遗传概念。这些活动结合了对本科和研究生阶段青年科学家的培训，并为高中生提供研究经验。参数化描述了一种行为，其中特定的跨同源相互作用导致减数遗传调控改变。这种行为可能是由小rna介导的，但其机制在很大程度上仍然未知。利用植物和花的颜色建立了一个模型系统来定义玉米发生参数化的分子机制。紫色植物t1基因的表达用可见色素进行监测。Pl1-Rhoades （Pl1-Rh）等位基因的强紫色状态可以改变为称为Pl'的较弱表达状态。当紫色类型（Pl/Pl）与弱色素植物（Pl/Pl’）交配时，只得到弱色素的后代——从这些后代中只有Pl’状态是性传播的；这是一个典型的参数化例子。正向遗传学鉴定了重要的顺式作用区域和15个位点，其功能需要维持Pl'状态的抑制（rmr）。先前的项目帮助鉴定了五种RMR蛋白在假定的小rna导向的DNA甲基化途径中起作用，并导致发现了一种共同转录抑制机制，该机制可维持通过参数化建立的减数遗传调控状态。本项目建立在这一新兴的机制理解基础上，结合正向遗传学、分子谱和突变分析，具体来说：1)通过谱分析转录、小RNA水平和参数化事件期间的胞嘧啶甲基化模式来测试参数化机制的工作模型；2)通过突变Pl1-Rh等位基因的结构特征，从功能上定义负责参数化行为的顺式作用序列。3)通过额外RMR位点的位置克隆发现控制参数的其他分子成分。培训和外展机会被纳入这些努力，以提供对基础真核遗传学和基因组生物学的更深入了解。该项目由分子和细胞生物科学部的遗传机制集群和综合生物系统部的植物基因组研究计划共同资助。

This project investigates a biological mechanism, called paramutation, which is responsible for turning selected genes "on" and "off" in highly predictable ways and for passing these "on" and "off" states to future generations. Similar to the well-known process of mutation, paramutation generates heritable biological variation that impacts evolutionary success. Unlike mutations, however, paramutations occur without changes to the DNA code. The project thus addresses an exception to an established rule of modern genetics by asking how regulatory information is altered and transmitted to offspring in the absence of DNA changes. The work will be carried out in corn, the preeminent model for studying the paramutation mechanism because the "on" and "off" states of genes responsible for plant color are visually tracked, controlled matings are simple to perform, and detailed multigenerational pedigrees can be generated and evaluated. A working model for paramutation has been proposed based on research and this project tests specific aspects of that model. The project will also identify novel molecules and chromosome structures responsible for the "on" and "off" switching during paramutations. Because paramutation occurs in both plants and animals, the results of this research could have broad biological impact, ranging from increased understanding of reproductive biology and genetics to development of novel strategies for improving agriculture or animal health. Educational materials such as colorful corn ears will be generated and made available through existing outreach programs for teaching both basic and advanced genetic concepts in the classroom. The activities integrate the training of young scientists at undergraduate and graduate levels as well as provide research experiences for high school students. Paramutation describes a behavior in which specific trans-homolog interactions result in meiotically-heritable regulatory alterations. This behavior is potentially mediated by small RNAs but the mechanism remain largely unknown. A model system was developed using plant and flower color to define the molecular mechanism of paramutation occurring in maize. Expression of the purple plant1 gene is monitored with visible pigments. A strong purple state of the Pl1-Rhoades (Pl1-Rh) allele can change to weaker expression states referred to as Pl'. When purple types (Pl/Pl) are mated with weakly pigmented plants (Pl/Pl'), only weakly pigmented progeny - from which only Pl' states are sexually transmitted - are obtained; this is a classic example of paramutation. Forward genetics identifies important cis-acting regions and fifteen loci whose functions are required to maintain repression (rmr) of Pl' states. Prior projects helped identify five RMR proteins acting in a presumed small RNA-directed DNA methylation pathway and led to the discovery of a co-transcriptional repression mechanism operating to maintain meiotically-heritable regulatory states established by paramutation. This project builds on this emerging mechanistic understanding by combining forward genetics, molecular profiling, and mutant analyses to specifically 1) test working models of the paramutation mechanism by profiling transcription, small RNA levels, and cytosine methylation patterns during paramutation events, 2) functionally define cis-acting sequences responsible for paramutation behaviors through structural characterizations of mutant Pl1-Rh alleles, and 3) discover additional molecular components governing paramutations through positional-based cloning of additional rmr loci. Training and outreach opportunities are integrated in these efforts to provide a greater understanding of fundamental eukaryotic genetics and genome biology.This project is jointly funded by the Genetics Mechanisms Cluster in the Division of Molecular and Cellular Biosciences and the Plant Genome Research Program in the Division of Integrative Organismal Systems.