多组学数据揭示棉花纤维发育的遗传学和表观遗传学基础
发布时间：2018-08-29 10:01
【摘要】：棉花是世界上重要的经济作物,棉花纤维是重要的天然纺织纤维。棉花纤维作为高度分化的单细胞,是研究细胞分化、细胞伸长和细胞壁合成的良好模型。本研究以组装和分析海岛棉基因组为基础,分析棉花纤维驯化历程和鉴定控制纤维品质性状形成的重要位点,并且分析纤维发育过程中的长链非编码RNA,进一步比较纤维和非纤维组织中可变剪接的差异,最后结合多组学数据,从表观遗传角度揭示纤维发育的动态过程。主要结果如下:1.海岛棉基因组测序和纤维细胞壁合成的重要基因家族分析本研究利用全基因组鸟枪法测序拼接四倍体海岛棉基因组。海岛棉基因组编码80876个基因,含有69%以上的重复序列。通过分析纤维素合酶(CesA)基因家族在棉花中的进化和各个成员在纤维发育过程中的表达模式,我们创造性地提出海岛棉纤维发育过程中CesA基因家族的接力模型,即At、Dt亚基因组合作调控四倍体棉花纤维发育。研究重点分析了果胶去甲酯化酶(PME)和果胶去甲酯化酶抑制基因家族(PMEI),发现PME的进化起源与细胞壁中果胶出现的时间一致。此外,研究发现这两个基因家族在高等植物中大量扩增,并且进化速率很快。我们推测PMEI结构域的起源可能是全基因组复制之后pro结构域的新功能化。这些结果促进了棉花基因组研究,加深了对植物细胞壁进化的理解。2.基因组重测序揭示了大量变异,这些变异是棉花驯化和顺式调控分化的基础本研究收集了352份陆地棉材料,包括半野生种和驯化种,构建了一个综合性的基因组变异图谱。研究鉴定了93个驯化选择区域,覆盖At亚基因组的74 Mb,Dt亚基因组的104 Mb,同时通过全基因组关联分析鉴定了19个与纤维品质性状相关的候选位点。结果表明长纤维的驯化与非对称的亚基因组选择相关。通过系统地分析DNase I酶切超敏感位点和三维基因组图谱,我们将一些可能存在的功能变异与基因转录联系起来,从而揭示了驯化对群体顺式调控元件分化的影响。本研究提出了一个重要作物中基因组的进化、调控和适应的新观点,为基于基因组的棉花遗传改良提供了丰富资源。3.棉花长链非编码RNA的鉴定和在纤维中的功能分析通过整合大量的转录组数据,本研究鉴定了30550个基因间区的长链非编码RNA(lincRNA)和4718个反义长链非编码RNA(lncNAT)位点。lncRNA的表达量比较低,并且大多数呈现出组织特异性表达模式。与蛋白质编码基因相比,lncRNA在基因区域的甲基化水平明显更高,并且其表达水平受基因区域甲基化影响较小。我们利用实验验证了一部分在棉花纤维发育起始期表达的lncRNA,发现这些lncRNA在纤维发育突变体和正常表型材料中差异表达,推测其可能与棉花纤维起始相关。最后,我们构建了共表达网络,对在纤维伸长和次生壁合成时期的lncRNA进行功能注释。4.棉花可变剪接图谱的建立及其复杂性和调控机制本研究利用单分子长读段转录本测序技术(Iso-Seq)对海岛棉进行转录组测序,开发了一套分析Iso-Seq数据的流程。我们从44968个基因中鉴定了176849个全长转录本,并更新了基因模型;同时,鉴定了15102个纤维特异的可变剪接事件,估计大约51.4%的亚基因组间的同源基因会通过可变剪接产生结构差异的转录本。进一步研究发现,同一个基因由可变剪接产生的转录本会受到miRNA的差异性调控。最后,我们发现核小体密度和DNA甲基化会在染色质水平定义外显子。本研究揭示了四倍体棉花中可变剪接的复杂性和新的调控机制,进一步增强了人们对多倍体植物中可变剪接的认识。5.棉花纤维发育动态表观组的建立和重要调控基因分析本研究建立了棉花纤维发育过程中的动态DNA甲基化组。随着纤维发育,DNA甲基化的比例会逐渐升高,呈现出与RNA介导的DNA甲基化相反的趋势。核小体定位分析表明发育中的纤维不断异染色质化,其可能与DNA甲基化的动态变化相关。绝大部分DNA高甲基化是由H3K9me2依赖的途径建立,与RNA介导的DNA甲基化不相关。通过分析四倍体棉花At和Dt亚组同源基因甲基化和表达量之间的关系,我们推测不同亚组基因的表观修饰差异可能会是基因偏向性表达的原因之一。最后,我们利用多组学数据的整合分析,揭示了DNA甲基化的动态变化可能会调控纤维细胞伸长和次生壁加厚时期活性氧含量的动态平衡。本研究首次绘制了棉花纤维发育过程中表观遗传修饰的动态图谱。
[Abstract]:Cotton is an important economic crop in the world, and cotton fiber is an important natural textile fiber. As a highly differentiated single cell, cotton fiber is a good model for studying cell differentiation, cell elongation and cell wall synthesis. The main results are as follows: 1. Sequencing of island cotton genome and the dynamic process of fiber development in terms of epigenetics. The tetraploid island cotton genome encodes 80876 genes with more than 69% repetitive sequences. The evolution of cellulose synthase (CesA) gene family in cotton and the expression patterns of each member during fiber development were analyzed. In this paper, we creatively proposed a model of the CesA gene family during fiber development in island cotton, i.e. at, Dt subgenomes cooperate to regulate fiber development in tetraploid cotton. We speculate that the origin of the PMEI domain may be a new functionalization of the pro domain after whole-genome replication. These results promote cotton genome research and deepen the understanding of plant cell wall evolution. 2. Genome re-sequencing revealed a large number of mutations, which were the basis of domestication and cis-regulation of cotton differentiation. This study collected 352 upland cotton materials, including semi-wild species and domesticated species, to construct a comprehensive genomic variation map. 93 domesticated selection regions covering 74 Mb and Dt subgenomes of the At subgenome were identified. 104 Mb. Nineteen candidate sites related to fiber quality traits were identified by genome-wide association analysis. The results showed that domestication of long fibers was associated with asymmetric subgenome selection. By systematically analyzing the DNase I digestion supersensitive sites and three-dimensional genome maps, we have identified some possible functional variations and gene transduction. This study proposes a new viewpoint on the evolution, regulation and adaptation of the genome in an important crop, which provides abundant resources for genome-based genetic improvement of cotton. 3. Identification of long-stranded non-coding RNA in cotton and functional analysis in fiber Integrating a large number of transcriptome data, we identified 30550 long-stranded non-coding RNA (lincRNA) and 4718 antisense long-stranded non-coding RNA (lncNAT) loci. The expression of lncRNA was relatively low, and most of them showed tissue-specific expression patterns. The expression level of some lncRNA expressed at the initial stage of cotton fiber development was verified by experiments. It was found that these lncRNA were differentially expressed in fiber development mutants and normal phenotypic materials. It was speculated that the expression level might be related to cotton fiber initiation. Establishment of cotton alternative splicing map and its complexity and regulation mechanism. In this study, Iso-Seq was used to sequence the transcripts of island cotton, and a procedure for analyzing Iso-Seq data was developed. 176,849 full-length transcripts were identified from 44,968 genes and the gene model was updated. Meanwhile, 1512 fiber-specific alternative splicing events were identified. It is estimated that about 51.4% of the homologous genes among subgenomes will produce transcripts with structural differences through alternative splicing. Further studies have shown that the same gene is produced by alternative splicing. Finally, we found that nucleosome density and DNA methylation define exons at the chromatin level. This study revealed the complexity of variable splicing in tetraploid cotton and new regulatory mechanisms, further enhancing the understanding of variable splicing in polyploid plants. 5. Cotton fiber hair Dynamic DNA methylation group was established during cotton fiber development. With the development of cotton fiber, the proportion of DNA methylation increased gradually, showing the opposite trend to RNA-mediated DNA methylation. Most of the DNA hypermethylation is established by H3K9me2-dependent pathway and is not related to RNA-mediated DNA methylation. Finally, the dynamic changes of DNA methylation may regulate the dynamic balance of reactive oxygen species (ROS) content during fiber elongation and secondary wall thickening. This study is the first time to map epigenetic modification in cotton fiber development.
【学位授予单位】：华中农业大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：S562
本文编号：2210865
[Abstract]:Cotton is an important economic crop in the world, and cotton fiber is an important natural textile fiber. As a highly differentiated single cell, cotton fiber is a good model for studying cell differentiation, cell elongation and cell wall synthesis. The main results are as follows: 1. Sequencing of island cotton genome and the dynamic process of fiber development in terms of epigenetics. The tetraploid island cotton genome encodes 80876 genes with more than 69% repetitive sequences. The evolution of cellulose synthase (CesA) gene family in cotton and the expression patterns of each member during fiber development were analyzed. In this paper, we creatively proposed a model of the CesA gene family during fiber development in island cotton, i.e. at, Dt subgenomes cooperate to regulate fiber development in tetraploid cotton. We speculate that the origin of the PMEI domain may be a new functionalization of the pro domain after whole-genome replication. These results promote cotton genome research and deepen the understanding of plant cell wall evolution. 2. Genome re-sequencing revealed a large number of mutations, which were the basis of domestication and cis-regulation of cotton differentiation. This study collected 352 upland cotton materials, including semi-wild species and domesticated species, to construct a comprehensive genomic variation map. 93 domesticated selection regions covering 74 Mb and Dt subgenomes of the At subgenome were identified. 104 Mb. Nineteen candidate sites related to fiber quality traits were identified by genome-wide association analysis. The results showed that domestication of long fibers was associated with asymmetric subgenome selection. By systematically analyzing the DNase I digestion supersensitive sites and three-dimensional genome maps, we have identified some possible functional variations and gene transduction. This study proposes a new viewpoint on the evolution, regulation and adaptation of the genome in an important crop, which provides abundant resources for genome-based genetic improvement of cotton. 3. Identification of long-stranded non-coding RNA in cotton and functional analysis in fiber Integrating a large number of transcriptome data, we identified 30550 long-stranded non-coding RNA (lincRNA) and 4718 antisense long-stranded non-coding RNA (lncNAT) loci. The expression of lncRNA was relatively low, and most of them showed tissue-specific expression patterns. The expression level of some lncRNA expressed at the initial stage of cotton fiber development was verified by experiments. It was found that these lncRNA were differentially expressed in fiber development mutants and normal phenotypic materials. It was speculated that the expression level might be related to cotton fiber initiation. Establishment of cotton alternative splicing map and its complexity and regulation mechanism. In this study, Iso-Seq was used to sequence the transcripts of island cotton, and a procedure for analyzing Iso-Seq data was developed. 176,849 full-length transcripts were identified from 44,968 genes and the gene model was updated. Meanwhile, 1512 fiber-specific alternative splicing events were identified. It is estimated that about 51.4% of the homologous genes among subgenomes will produce transcripts with structural differences through alternative splicing. Further studies have shown that the same gene is produced by alternative splicing. Finally, we found that nucleosome density and DNA methylation define exons at the chromatin level. This study revealed the complexity of variable splicing in tetraploid cotton and new regulatory mechanisms, further enhancing the understanding of variable splicing in polyploid plants. 5. Cotton fiber hair Dynamic DNA methylation group was established during cotton fiber development. With the development of cotton fiber, the proportion of DNA methylation increased gradually, showing the opposite trend to RNA-mediated DNA methylation. Most of the DNA hypermethylation is established by H3K9me2-dependent pathway and is not related to RNA-mediated DNA methylation. Finally, the dynamic changes of DNA methylation may regulate the dynamic balance of reactive oxygen species (ROS) content during fiber elongation and secondary wall thickening. This study is the first time to map epigenetic modification in cotton fiber development.
【学位授予单位】：华中农业大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：S562
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