种子发育相关B3基因的起源和进化

【作者】 李杨；

【导师】 杨继；

【作者基本信息】 复旦大学 ， 生态学， 2011， 硕士

【摘要】 种子形成(seed formation)是陆生植物进化的关键创新(key innovation)之一,也是种子植物广泛适应不同陆地环境并发展成为陆地植被主要类群的基础。从系统发育的角度看,种子的形成实际是由于在高等植物生活史中、在胚胎发生的后期插入了一个成熟阶段所致。B3类转录因子在植物基因组中形成一个超家族,包括：LAV家族、ARF家族、RAV家族和REM家族,其中LEC2、FUSCA3和ABI3/VP1隶属于LAV家族、AFL亚家族,它们彼此互作形成一个调控网络,在种子植物基因组中调节种子成熟发育过程。但在系统演化过程中,以LEC1、LEC2、FUS3和ABI3为主调控因子的种子成熟调控模块何时形成?是否伴随着种子植物的分化而发生?不同种子植物(包括裸子植物、单子叶植物、双子叶植物)种子成熟发育的分子调控机制是否一致?是否基于相同主调控基因的作用?种子性状如何从一个生活史中不具休眠胚阶段的祖先产生?这些都还是目前悬而未决的问题。本项研究旨在：(1)以种子发育主控基因的进化分析为主线,充分利用现有的植物全基因组信息,运用比较基因组和分子进化分析手段,对LEC2、FUS3、ABI3和其它种子发育相关转录调控基因在不同类群植物基因组中的分布状况、变异式样和进化趋势进行全面的比较和分析；(2)利用分子生物学手段扩增目前尚无基因组信息的裸子植物AFL亚家族相关基因,以帮助探讨种子成熟发育调控网络的进化时间和机制。研究结果表明：(1)AFL亚家族成员在陆生植物基因组中广泛存在。其中,ABI3IVP1基因与绿藻的一个单拷贝B3基因有较高的同源性,并分布于所有陆生植物类群的基因组中；FUSCA3伴随裸子植物的分化而产生,主要分布于种子植物基因组中；而日前证据显示,LEC2主要分布在双子叶植物的基因组中,在禾本科植物中存在另一类与AFL业家族成员序列相似程度较高的基因——-IDEF-like基因。因此,AFL家族成员是伴随着陆生植物不同类群的分化而逐步产生的。(2)AB13/VPI是AFL亚家族中最早出现的成员,其蛋白序列中含有多个保守的结构域或功能域,伴随FUS3和LEC2的分化,保守结构域/功能域的数目表现出递减的趋势,且FUS3在禾本科植物与双子叶植物中分化出了各自特异的C端保守区,表明在进化过程中AFL亚家族成员经历了不同类型和不同程度的结构和功能特化。(3)从基因外显子-内含子结构与数目看,AFL亚家族成员在进化历程中经历了频繁的内含子获得事件,根据基因外显子-内含子结构推测,小立碗藓ABI3的一个拷贝是其他所有维管植物AFL亚家族成员的祖先。(4)运用PCR和[AIL-PCR方法从松杉纲植物和银杏中成功扩增得到ABI3及FUS3的同源序列,支持了基于生物信息学和比较基因组学的研究结果。目前普遍认为,ABI3、FUS3和LEC2是种子形成和发育的主调控因子,但本项研究结果它们并不是伴随着种子植物的分化而同步产生的。ABI3在种子植物发生以前就已经存在,在种子性状进化过程中被招募到新的基因调控网络中发挥新的功能；而LEC2则是后期伴随着被子植物(或双子叶植物)的分化而产生的。因此,该项研究结果对从进化发育生物学(Evo-Devo)角度探讨种子性状系统发生的分子遗传学基础具备重要参考价值。更多还原

【Abstract】 Seed formation is one of the key innovations during land plant evolution, which endowed plants with higher adaptability toward unfavorable environment and strengthened the ability of dispersal and competition. The process of seed development could be divided into two phases, i.e. the early embryo morphogenesis phase and the late maturation phase. During the late maturation phase, the embryo accumulates various storage compounds and acquires desiccation tolerance, and eventually enters dormancy.B3 genes encode plant-specific transcription factors, which play a variety of roles in plant growth and development. Proteins encoded by B3 genes vary in sequence similarity and domain structure but all contain a highly conserved B3 domain, forming the B3 superfamily. This superfamily consists of several distinct families, including the LAV, ARF, RAV and REM families, with a few subfamilies to be identified in each family. The AFL subfamily belongs to the LAV family, containing three members:ABSCISIC ACID INSENSITIVE 3(ABI3), FUSCA 3 (FUS3) and LEAFY COTYLEDON 2 (LEC2). The three members of the AFL subfamily are considered master genes in controlling seed development, especially for seed maturation. It remains unclear, however, whether all these genes are necessary and sufficient for induction of the maturation phase of seed development for all seed plants. The questions such as whether these genes appeared at the same time as the origin of the seed plants and how they originated remain unknown.In this study, genome-wide screens for the AFL genes were performed based on currently available plant genome sequences. By conducting a comprehensive analysis of the distribution patterns, gene structures and phylogenetic relationships of ABI3, FUS3 and LEC2, we intend to address questions, such as when these genes originated, how they duplicated and diverged during evolution, and whether all these genes are indispensable components of the regulatory network involved in the control of seed development for all seed plants. We also cloned and characterized the AFL homologous genes from several gymnosperm plants, for which the complete genome sequences were not available.The results showed that the AFL subfamily genes are widely distributed in land plant genomes, and that the AFL genes underwent a series of independent duplication and diversification during land plant evolution. The duplicate ABI3 genes of non-seed plants seem to have structurally and functionally diverged prior to the diversification of seed plants, leading to the reduction of the numbers of ABI3 in the seed plant genomes and giving rise to the appearance of the FUS3 gene in gymnosperms. In accompany with the divergence of gymnosperm and angiosperm, the FUS3 gene experienced a new round of duplication with subsequent diversification through neo-and/or sub-functionalization. As a result, one copy retained the original function of FUS3 in both gymnosperm and angiosperm genomes and the other copy diverged into the ancestors of LEC2 and IDEF-like. Present data suggested that the LEC2 gene appeared initially and limitedly distributed in the dicot genomes, while the functionally diverged IDEF-like gene limited to the monocot genomes. Therefore, the ABI3, FUS3 and LEC2 genes have appeared in the land plant genomes in a stepwise pattern. There is a clear trend toward the reduction of the conserved domain contained in ABI3, FUS3 and LEC2. Compared to the wide range of expression pattern of ABI3, FUS3 and LEC2 exhibit restricted spatial and temporal expression. Functional diversification of the AFL genes seems to correlate with the variation in domain structure.ABI3, FUS3 and LEC2 are currently considered the master regulatory genes for seed maturation. Based on the currently available data, the LEC2 genes are only distributed in the dicot genomes. It is thus doubtful whether all the ABI3, FUS3 and LEC2 genes are indispensable components of the regulatory network controlling seed development for all seed plants. To answer this question, we need more complete genome sequences, especially the genomic information of gymnosperms. Meanwhile, the evolutionary relationships between LEC2 and IDEF remain to be further assessed. These two genes probably descended from different ancestors that appeared at the early stage of angiosperm diversification, with the LEC2 and IDEF homologs sequences to be lost in the monocot and dicot genomes, respectively. But we also cannot exclude the possibility that IDEF was derived from LEC2 through duplication and subsequent diversification during the monocots-dicots split. To investigate when these genes originated and how they diverged during evolution will provide crucial insights into the molecular mechanisms controlling seed formation and evolution.更多还原