Epigenetics and hidden heritability in tomato

真核生物基因组的信息内容主要基于DNA序列，但还有其他层依赖于DNA中胞嘧啶残基的甲基化和与染色体中DNA结合的组蛋白的修饰。这种表观遗传信息的一个主要功能是防止DNA寄生虫——转座因子——在基因组中定居并具有诱变特性。因此，表观遗传学是宿主基因组和这些DNA寄生虫之间军备竞赛的一部分，但就像所有军备竞赛一样，这种动态并不简单。如果寄生虫具有增加宿主健康的特征或作用，宿主甚至可能受益，相反，如果宿主的防御系统减少寄生造成的损害，寄生虫也可能受益（寄生虫依赖于宿主的健康）。这种复杂动态关系的证据来自植物DNA甲基化突变体的表型，其中DNA甲基化在整个基因组中丢失。这不仅会导致转座子防御功能的丧失，还会破坏植株的生长发育和不育性。然而，基因组局部DNA甲基化的缺失可能会产生更具体的影响，包括一些有益的影响。这些观察结果提出了一种假设，即表观遗传学可能解释了作物育种者所熟知的隐性遗传性。隐藏的遗传能力最好地用数量性状来说明，因为植物之间的许多可遗传变异不能与遗传标记联系起来。在某些情况下，这种隐性遗传或缺失遗传可以解释变异的很大一部分。对植物表观遗传学的分子认识大多来自模式植物拟南芥。广泛的资源促进了对这种植物的实验，并使人们对各种表观遗传机制有了很好的理解。然而，这种模式物种的快速进展部分是因为基因组小，与大多数其他物种（包括作物）相比，转座因子少得多。因此，这种减少的表观基因组的扰动对植物的生长和发育有相当轻微的影响。在具有较大的、富含转座子的基因组的植物中，转座子对生长发育的影响更深远，这与表观遗传转座子防御与基因组正常功能重叠一致。不幸的是，使用标准的DNA甲基化突变体，植物可能是不育的，并且难以在实验中使用。为了解决这个问题，我们在番茄中开发了一种DNA甲基化突变的方法，这种突变会对表观基因组造成有限的破坏。在我们的准备工作中，我们已经分离并鉴定了DNA甲基化基因CMT3a的突变体。部分效应允许恢复可育突变植株，以便我们可以实施cmt3a依赖性表观基因组及其影响的详细分析。在项目的第一部分，我们将全面表征cmt3a依赖性表观基因组及其对基因表达的影响。在我们的初步证据中，我们表明基因组的一些区域在突变计划中失去了DNA甲基化，并且在回交到野生型植物的后代中保持了这种低甲基化状态，而其他区域则恢复了正常的DNA甲基化水平。将这些发现与对cmt3a及其后代基因表达的分析相结合，将确定基因组中与表观基因组遗传能力相关的特征及其对基因表达的影响。然后，我们将通过使用改良的CRISPR系统对表观基因组进行靶向修饰来测试这些相关性。该表观基因组靶向修饰系统也将作为一种表观遗传修饰系统在作物植物改良方面进行探索。

The information content of eukaryotic genomes is based primarily on the DNA sequence but there are additional layers that are dependent on methylation of cytosine residues in the DNA and modification of the histone proteins that bind to DNA in the chromosomes. A primary function of this epigenetic information is to protect against DNA parasites - transposable elements - that colonise genomes and have mutagenic properties. Epigenetics, therefore, is part of the arms race between the host genomes and these DNA parasites but, like all arms races,the dynamics are not simple. The host may even benefit if a parasite has features or effects that increase the host's fitness and, conversely, the parasite may also benefit if the defense systems of the host reduce the damage caused by parasitism (the parasite is dependent on the well-being of the host). Evidence of this complex dynamic relationship is from the phenotype of DNA methylation mutants in plants in which DNA methylation is lost throughout the genome. There is not only loss of transposon defense but additionally disrupted growth and development of the plant and sterility. Loss of the DNA methylation in localized parts of the genome, however, may have more specific effects including some that are beneficial. These observations prompt the hypothesis that epigenetics may account for the hidden heritability that is well known to breeders of crops. Hidden heritability is best illustrated with quantitative traits for which much of the heritable variation between plants cannot be linked to genetic markers. In some instances this hidden heritability or missing inheritance can account for a substantial part of the variation. Much of the molecular understanding of epigenetics in plants is from the model plant Arabidopsis. Extensive resources that have facilitated experimentation with this plant and it has led to good understanding of the various epigenetic mechanisms. The rapid progress with this model species, however, is partly because the genome is small and has many fewer transposable elements than most other species, including crops. Perturbation of this reduced epigenome, therefore, has rather mild effects on the growth and development of the plant. In plants with larger, transposon-rich genomes there are more profound effects on growth and development consistent with the epigenetic transposon defense overlapping with the normal function of the genome. Unfortunately, with the standard set of DNA methylation mutants, the plants may be sterile and be difficult to use experimentally. To address this problem we have developed an approach in tomato with a DNA methylation mutant that causes limited disruption of the epigenome. In our preparatory work we have isolated and characterized such a mutant in the DNA methylation gene CMT3a. The partial effect allows recovery of fertile mutant plants so that we can implement a detailed analysis of the CMT3a-dependent epigenome and its effects. In the first part of the proposed project we will fully characterize the CMT3a-dependent epigenome and its influence on gene expression. In our preliminary evidence we show that some regions of the genome lose their DNA methylation in the mutant plans and that they retain this hypomethylated state in backcrossed progeny to wild type plants whereas other regions regain the normal levels of DNA methylation. Integrating these findings with analyses of gene expression in cmt3a and its progeny will identify features in the genome that correlate with heritability of the epigenome and its effects on gene expression. We will then test these correlations by targeted modification of the epigenome using a modified CRISPR system. This system for targeted modification of the epigenome will also be explored as a system for epigenetic modification in crop plant improvement.