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题名: 玻璃海鞘(Ciona intestinalis)快速局域适应的基因组学效应
作者: 林亚萍
学位类别: 博士
答辩日期: 2016-06
授予单位: 中国科学院研究生院
授予地点: 北京
导师: 战爱斌
关键词: 海鞘,生物入侵,微进化,基因组扫描,自然选择 ; Ascidian, Biological invasion, Microevolution, Genome scan, Natural selection
其他题名: Genomic signatures of rapid local adaptation in the model invasive ascidian, Ciona intestinalis
学位专业: 环境科学
中文摘要:     局域适应是入侵生物适应新栖息地环境的重要微进化过程之一。玻璃海鞘作为入侵生物的模式物种,对温度和盐度具有很强的适应性,是解析全球气候变化背景下的微进化过程的理想材料。虽然快速局域适应在入侵生物中普遍存在,但是哪些基因参与了快速局域适应?快速局域适应在生物基因组上留下哪些印记?快速局域适应的环境驱动因子有哪些?诸多问题亟待解决。本研究以玻璃海鞘为研究对象,在全基因组水平上探讨了玻璃海鞘局域适应的遗传本质。

1. 全基因组水平玻璃海鞘复合种微卫星标记的开发与评价
      本研究通过公用数据库筛选得到全基因组水平的分子标记。通过 JGI 数据库我们得到684,393 条玻璃海鞘EST 序列,软件查找得到4,654 个微卫星,其中2,126 个微卫星能成功定位到玻璃海鞘染色体上。基于微卫星的功能注释与其在染色体上的分布情况,我们选择了545 个具有代表性的微卫星进行扩增与多态性评价。在玻璃海鞘A 种的两个群体(AM(可能是本地群体)和入侵群体SA)中成功扩增得到218 个具有多态性的微卫星标记。多态性评价结果显示,扩增得到的等位基因数目在AM 和SA 群体中分别从2~11 个和2~10 个不等,期望杂合度的范围分别为0~0.833 和0.021~0.818,观测杂合度的范围分别为0~0.879 和0.031~0.845。25.8%(60 个)的微卫星在玻璃海鞘B 种中成功扩增并具有多态性。

2. 玻璃海鞘自然群体多样性及其遗传分化研究
      已有的研究认为,玻璃海鞘全球群体之间具有很高的遗传同质性。为得到玻璃海鞘更为精细与全面的群体遗传结构,本研究利用覆盖全基因组的转录组微卫星,和经历不同选择类型的微卫星分别对5 个玻璃海鞘群体进行群体遗传结构分析。结果显示,玻璃海鞘群体之间存在很强的遗传分化,两两群体间的FST 值在0.0169~0.2041 之间变化。其中地中海群体(AM 和BL)、大西洋群体(SA)与太平洋群体(NMF 和GAP)之间的遗传分化最大,FST 在0.1834~0.2041之间。3D-FCA 和贝叶斯聚类分析得到一致的结果,两个地中海群体和一个大西洋群体聚成了一簇,其余两个太平洋群体聚成了另外一个簇。分别用不同受
选择类型(中性位点、正向选择和平衡选择位点)位点进行群体遗传结构比较分析发现,正向选择位点较中性位点得到更为清晰的遗传结构。说明自然选择作用导致了玻璃海鞘群体之间的适应性分化。比较已有的基因组微卫星的研究结果,我们证实了转录组微卫星较基因组微卫星对群体遗传结构具有更强的区分力,尤其是在基因流较强的海洋生物中。

3. 全基因组扫描自然选择在玻璃海鞘基因组上的印记
    自然选择被认为是适应性进化的主要动力。本研究通过全基因组扫描,结合选择性扫除原理筛查玻璃海鞘基因组自然选择的信号,从基因组水平阐述玻璃海鞘局域适应的本质。应用F 异常值检验,基于玻璃海鞘基因组物理图谱,采用LOSITAN,BAYESCAN 和ARLEQUIN 三种方法,共得到了42(27.6%)个参与玻璃海鞘快速局域适应过程的基因组区域。19(12.5%)个区域具有正向选择信号,其中12 个独立分布于9 条染色体上。在两两群体比较分析中,所有受选择位点均涉及NMF 群体或者GAP 群体,说明入侵群体,尤其是太平洋群体(NMF 和GAP)受到的选择压力明显高于地中海群体(可能是本地群体,AM 和BL)。根据受选择位点相邻位点的受选择情况,发现玻璃海鞘基因组选择性扫除的区域较小,最小的选择性扫除区域小于8.9 kb。由此推测,玻璃海鞘的的自然选择作用很有可能基于固有遗传变异。通过受选择位点与环境因子的相关分析,得到2 个受选择位点与盐度相关。因此,盐度差异很有可能是玻璃海鞘产生局域适应的重要环境驱动因子之一。

4. 玻璃海鞘快速局域适应候选基因研究
    通过分析受选择位点上下游的基因,我们共得到 115 个参与快速适应进化的候选基因。包括钙调蛋白4 基因、细胞程序性死亡蛋白2 基因、ciliogenesis-associated TTC17-interacting protein 基因、动力蛋白重链2 基因和血蓝蛋白基因等。已有的研究证实这些基因在渗透压(包括钙离子浓度)调节、精子活力、信号通路调节、细胞的增殖与凋亡、氧气输送和有丝分裂等过程中都发挥着重要作用。因此很有可能参与到玻璃海鞘的快速适应进化中。这些选
择区域重点基因的分析与遴选为今后的研究工作奠定了基础。通过解析这些基因内部及上、下游区域在DNA 水平上在来源于不同地域的群体中发生变化,阐明群体快速适应迅速变化的环境及栖息地转换的遗传学本质。
    本研究阐述了自然选择作用驱动玻璃海鞘群体的适应性分化,探讨了自然选择在物种基因上的印记与对基因组结构的影响,初步揭示了驱动自然选择的环境梯度因子。在基因组水平为阐明玻璃海鞘对入侵地变化环境(水温和盐度等)的适应性进化过程提供了理论依据,为自然选择导致的物种适应性进化过程这一科学问题提供重要例证。
英文摘要:     Local adaptation is one of the important microevolution in invasive species as a response to the changing environment of the invaded habitat. Ciona intestinalis, as a unique model for studying biological invasion, could rapidly adapt to varying habitat environments, typically habitats with dramatically different temperature and salinity. Thus, C. intestinalis provides an excellent model for the study of microevolution associated with changing environments which are increasingly affected by global climate change. Although rapid local adaptation is believed to frequently occur in invasive species, it remains largely unknown that how rapid environmental change and invasive behavior affect genomes. For example, which genes are involved in the rapid local adaptation? Which environment variables determining the fitness of these genes? We adopted C. intestinalis as a system to detect genomic signatures of rapid
local adaptation and to investigate the genetic mechanism of local adaptation of C.
intestinalis

1. Genomewide gene-associated microsatellite markers for C. intestinalis
    In this study, we characterized a large set of genome-wide gene-associated microsatellite markers for C. intestinalis spA. We downloaded all 684,393 EST sequences of C. intestinalis spA from the Joint Genome Institute (JGI) website. Bioinformatic analysis identified 4,654 microsatellites from expressed sequence tags (ESTs), 2,126 of which successfully assigned to chromosomes were selected for further analysis. Based on the distribution evenness on chromosomes, function annotation and suitability for primer design, we chose 545 candidate microsatellites for further characterization. After amplification validation and variation assessment, 218 loci were polymorphic in at least one of the two populations collected from the coast of Arenys de Mar, Spain (N = 24 - 48) and Cape Town, South Africa (N = 24 - 33). The number of alleles, observed heterozygosity and expected heterozygosity ranged from two to 11, 0 to 0.833 and 0.021 to 0.818, and from two to 10, 0 to 0.879 and 0.031 to 0.845 for the Spanish and African populations, respectively. When all
microsatellites were tested for cross-species utility, only 60 loci (25.8%) could be successfully amplified and all loci were polymorphic in C. intestinalis spB. A high level of genome-wide polymorphism is likely responsible for the low transferability. The large set of microsatellite markers characterized here is expected to provide a useful genome-wide resource for evolutionary studies using C. intestinalis as a model.

2. Genetic diversity and differentiation of C. intestinalis in wild populations
    Previous studies found a very high level of genetic homogeneity and lack of geographical differentiation across a wide geographical area among populations of C. intestinalis. To get finer and more comprehensive coverage of the patterns of genetic structure, we use genome-wide gene-associated microsatellite markers and different marker categories (viz. neutral loci, loci under directional selection and loci under balancing selection) to investigate the population genetic structure of C. intestinalis.
      We first genotyped five global populations that were representative of the environmental conditions (seawater temperature and salinity) found across the likely native and introduced range of the studied species. This analysis showed significant population genetic differentiation among populations at the genomic level. Pairwise FST values ranged from 0.0169 between AM and BL to 0.2041 between AM and NMF. Populations from the Pacific coast (NMF and GAP) were highly differentiated from those collected from the Mediterranean Sea and Atlantic coast (AM, BL and SA; FST = 0.1834 - 0.2041). 3D-FCA and Bayesian clustering analysis have produced inconsistent results that showed a perfect separation of the samples into two groups -group 1 from Pacific coast (NMF and GAP) and group 2 from the Mediterranean Sea and Atlantic coast (AM, BL and SA). Moreover, all these directionally selected loci provided finer population structure than the neutral markers. Our results suggest that divergent selection maintains adaptive differentiation despite the homogenizing effect of artificial dispersal in C. intestinalis. and demonstrated that the
gene-associated marker provided improved resolution in respect to population structure, especially for species that inhabit high gene flow environments, such as marine invasive species.

3. Genome-wide scans for signatures of natural selection in C. intestinalis
      Natural selection is one of the most important drivers for adaptive evolution.We conducted genome-wide scans combined with hitchhiking map analysis for signatures of natural selection and thus illuminated the genetic mechanism of local adaptation of C. intestinalis. After combining results from the three theoretical approaches (i.e., Beaumont & Nichols’s (1996) method, hierarchical method and Bayescan method) based on the FST-outlier test for modeling neutral loci, a total of 42 (27.6%) loci were identified under directional or balancing selection, 19 (12.5%) and 23 (15.1%) of which were candidates for directional selection and balancing selection, respectively. Of the 19 candidates of directional selection loci, 12 were located in nine different chromosomes likely represented 12 distinct selective sweep regions on chromosomes, of which we inferred that the smallest sweep region was less than 8.9 kb. Therefore, we speculated that several soft selective sweeps were acted upon the genomic regions of C. intestinalis (i.e. selection acted on standing genetic variation). In pairwise analysis, all outlier loci were associated with NMF
and/or GAP which were likely to experience stronger selection pressure than populations from likely native regions (AM and BL). Moreover, two of the selected loci (Cin54 and Cin35) showed significant correlation with local salinity (P < 0.05) which were likely to be involved in adaptation to changing environments in C. intestinalis. We speculated that salinity gradient is one of the most important drivers for local adaptation of C. intestinalis.

4. Candidate genes for local adaptation in C. intestinalis
      We analyzed the selection windows along chromosomes and found 115 genes in regions under selective sweeps, including physiologically important genes such as calmodulin-like protein 4, ciliogenesis-associated TTC17-interacting protein-like,dynein heavy chain 2, programmed cell death protein 2 and the gene hemocyanin G-type, units Oda to Odg-like. These genes were identified important function of osmotic regulation, sperm motility, diverse signaling pathways regulation, oxygen transport, cell growth and death and mitosis which were likely to be involved in adaptation in C. intestinalis. Taken together, we found genomic regions under selection associated with local adaptation in a highly invasive species, and identified testable candidate genes for future analyses of evolutionary responses to rapid environmental change.
      Our study demonstrated adaptive differentiation driven by divergent selection,uncovered the genomic signatures of natural selection and revealed the environment gradient determining the fitness in C. intestinalis. These findings provide a good example for understanding adaptive evolution of invasion species driven by natural selection.
内容类型: 学位论文
URI标识: http://ir.rcees.ac.cn/handle/311016/36891
Appears in Collections:中科院环境生物技术重点实验室_学位论文

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Recommended Citation:
林亚萍. 玻璃海鞘(Ciona intestinalis)快速局域适应的基因组学效应[D]. 北京. 中国科学院研究生院. 2016.
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