拟南芥和蓖麻种子特异表达基因的表观调控研究

	拟南芥和蓖麻种子特异表达基因的表观调控研究
	吴迪
导师	刘爱忠
关键词	拟南芥，蓖麻，种子特异基因，DNA甲基化谷，组蛋白修饰 Arabidopsis, Castor bean, Seed specific genes, DNA methylation valley, Histone modification
摘要	种子是植物繁衍的载体，也是人类重要的食物来源及工业生产的原料。解析种子生长发育的分子调控机理一直是植物学、农学和种子生物学研究的核心内容之一。种子的正常生长发育受到了一系列种子特异表达基因组成的复杂网络的调控，解析这些基因在种子中特异表达形成的机制将为理解种子发育过程及优质种子作物选育的工作提供重要的理论及实践意义。近年来，尽管研究人员通过遗传学、分子生物学的手段鉴定到了大量影响种子生长发育的关键调控因子，阐明了许多关键基因对种子发育的分子调控机制，然而关于这些种子特异基因本身是如何在种子中被激活，从而参与种子发育调控的分子机制尚不明确。本研究以拟南芥及蓖麻两种不同类型的种子为研究对象，采集了拟南芥及蓖麻不同发育阶段的种子及不同的营养组织，进行了转录组测序、DNA甲基化测序及不同组蛋白标记的ChIP测序，并进行了多组学的关联分析，揭示了基因组表观特征DNA甲基化谷（DMV）中发生的组蛋白修饰重排对种子发育和储藏物质累积关键基因的调控机理。主要结果如下： 1. 利用多组织RNA-seq测序数据获得的基因表达谱，我们在拟南芥与蓖麻中分别鉴定了2625和1162个种子特异表达基因。根据基因在种子发育不同阶段的表达规律，将这些基因分为了三种表达模式，基因功能富集分析结果表明，三种表达模式下的基因分别与种子早期发育、中期种子储藏物质累积和后期种子脱水成熟密切相关。拟南芥与蓖麻中有212个共同的种子特异表达基因，其中包括LEC1、WRI1、ABI3及FUS3等调控种子发育的核心转录因子，表明这些基因在植物中对种子发育的调控作用是保守的。 2. 我们发现种子特异基因的DNA甲基化水平在不同组织中均低于组成型表达基因，表明DNA甲基化不直接调控种子特异基因的表达。进一步通过全基因DNA甲基化测序和分析，发现了拟南芥及蓖麻基因组上存在大量的DNA低甲基化区域，占了拟南芥基因组的56%和蓖麻基因组的33%。这些DMV在不同的组织及发育阶段均高度保守。我们在拟南芥及蓖麻中分别鉴定到11687及13800个DMV基因，发现DMV基因具有明显的组织特异表达的趋势，我们所鉴定的种子特异基因超过60%都位于DMV中，包括种子发育关键转录因子LEC1、LEC2、WOX2、DOG1等，表明DNA低甲基化是种子特异基因的重要表观特征。我们发现拟南芥及蓖麻之间共有4083个保守的DMV基因，对保守DMV基因的功能富集分析发现，DMV基因主要参与了转录调控过程，且在调控种子发育、调控分生组织干细胞特性及器官发育等植物发育过程中有极其重要的作用，暗示DMV基因与保守的植物发育过程密切相关。 3. 结合不同组蛋白修饰的ChIP-seq分析，我们发现绝大多数组蛋白修饰发生在DMV区域。通过比较位于DMV的种子特异基因在叶片及胚乳间的组蛋白修饰差异，我们发现在叶片中，种子特异基因被抑制型组蛋白修饰（H3K27me3）所占据，而在种子中，抑制型组蛋白修饰在种子特异基因上的富集减少，伴随着激活型组蛋白修饰（H3K36me3、H3K9ac、H3K27ac和H3K4me3）的富集增加，从而促进了种子特异基因的激活；而组成型表达基因在叶片及胚乳中的组蛋白修饰没有显著差异。我们发现一些远端DMV上有H3K27ac的富集，具有增强子的特征，双荧光素酶报告实验证明了这些远端DMV可以激活下游基因的表达。本研究首次揭示了植物DMV中发生的组蛋白修饰重排，对种子特异基因在种子中的激活有重要的调控作用，为全面理解种子发育和储藏和物质累积的分子调控机制提供了新的认识，具有重要的理论和实践意义。; Seeds are the link between plant vegetative and reproductive growth, also an important source of food for humans and materials for industrial production. Unraveling the molecular regulatory mechanisms of seed growth and development are essential for both plant biology, agronomy, and seed biology research. Seed growth and development is regulated by a complex regulatory network of seed-specific expressed genes. Elucidating the seed-specific expression mechanism of these genes will provide important theoretical and practical significance for understanding the basic seed development process and improving the crops breeding for high quality seed. In recent years, a growing number of studies have identified the key regulators that affect seed growth and development and uncovered the molecular regulatory mechanisms of these genes on seed development through genetic and molecular biological approaches, but how these genes are expressed in a seed?specific manner, thus involved in the regulation of seed development are not clear. In this study, we select Arabidopsis and castor bean represent two different seed types material. We collect seeds form different developmental stages together with various vegetative tissues for transcriptomic sequencing, DNA methylation sequencing and ChIP sequencing with different histone marks, and conducted a multi-omics association analysis with the datasets generated from different high-throughput sequencing. Our research revealed the regulatory mechanism of histone modification reconfiguration occurring in DNA methylation valley (DMV) on genes critical for seed development and storage compounds accumulation. The main results are as follows: 1. Based on gene expression profiles obtained from RNA-seq data, we identified 2625 and 1162 seed-specific genes in Arabidopsis and castor, respectively. Seed-specific genes were subsequently classified into three expression patterns according to the expression level in different seed development stages, functional enrichment analysis showed that genes from three expression patterns were closely related to early seed development, mid-term seed storage compounds accumulation, and late seed dehydration and maturation, respectively. There were 212 seed-specific genes common to both Arabidopsis and castor, including LEC1, WRI1, ABI3 and FUS3, which are core transcription factors regulating seed development, indicating that the regulatory roles of these genes on seed development are conserved in plants. 2. We found that DNA methylation level of seed-specific genes was lower than that of constitutive genes in different tissues, indicating that DNA methylation does not regulate the expression of seed-specific genes directly. Whole genome DNA methylation sequencing revealed that DNA hypomethylated regions were widely distributed in both Arabidopsis and castor genome, accounting for 56% of Arabidopsis and 33% of castor genomes. DMVs are highly conserved in different tissues and seed development
语种	中文
	2022-05
学位授予单位	中国科学院大学
文献类型	学位论文
条目标识符	http://ir.kib.ac.cn/handle/151853/75131
专题	昆明植物所硕博研究生毕业学位论文
推荐引用方式 GB/T 7714	吴迪. 拟南芥和蓖麻种子特异表达基因的表观调控研究[D]. 中国科学院大学,2022.

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