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题名	新外显子的起源与功能及短同源序列介导转录滑动产生嵌合RNA 的研究
作者	李昕
学位类别	博士
答辩日期	2008-06
授予单位	中国科学院研究生院
授予地点	北京
导师	王文
关键词	新外显子 快速进化 正选择 嵌合RNA 反式剪切 转录滑动
其他题名	Origin and function of new exons and generation of chimeric RNAs mediated by short homologous sequences
学位专业	动物学
中文摘要	新外显子的起源是一种重要的增加转录组和蛋白质组多样性的分子机制。对于新外显子及其父本基因的进化和功能特征方面还有很多重要的问题有待于解决。本研究首先在全基因组水平上鉴定在人和小鼠中产生的新外显子，随后对这些外显子及其父本基因作进化和功能上的分析。我们发现新外显子倾向于位于基因的UTR区域，尤其是5’ UTR区域，这表明可能有些新外显子的出现与基因的表达调控相关。我们还发现，产生新外显子的基因具有较高的组织表达特异性，其基因功能倾向于细胞调控和与外界环境相互作用。通过对外群中直系同源基因的分析，我们的结果表明进化速率较高的基因更容易获得新的外显子，纠正了先前认为的获得新外显子会加速基因进化速率的看法。 我们对哺乳类CDYL基因家族中产生的新外显子进行了具体的进化分析和功能研究。我们的结果表明CDYL基因在哺乳类分化前在原先的基因上游区域获得了一个新的启动子和三个新的外显子。随后在哺乳动物各个支系的分化中，CDYL基因在小鼠，狗和人中分别独立的进化出一个新的外显子。同源比对的结果表明，这些新外显子是通过内含子序列的外显子化这一分子机制产生。近缘物种间的进化速率的计算结果表明这些新产生的外显子具有快速进化的模式，并且其快速进化可能是由正选择所驱动。在人中，多种突变包括新外显子的获得，启动子的改变，选择性剪切的发生使得人的CDYL基因获得了一种新的编码更长蛋白质的剪切体。在人Hela细胞系中的实验表明，新产生的蛋白质与原有的蛋白质相比都具有显著的转录抑制活性，但新的蛋白质的转录抑制活性较弱，且两者之间存在相互干扰的关系。这一结果表明通过新外显子的获得产生的新的蛋白质可以丰富原有的基因表达调控体系，使得生物体的调控网络更加精确。 嵌合RNA通常认为是由来源于不同的pre-mRNA的外显子通过反式剪切连接在一起形成的。这一现象在包括多种动物和植物中被广泛的报道。我们的研究首先通过大规模表达序列（ESTs）的搜索，在酵母，果蝇，小鼠和人中鉴定到了大量的嵌合RNA。这一结果表明形成嵌合RNA在真核生物中是一种普遍的生物学过程，是一种重要的增加转录组和蛋白质组的多样性的分子机制。对嵌合RNA的序列分析表明，仅有<20%的嵌合RNA在接合处可以找到典型的剪切位点GU-AG，可以用经典的反式剪切模型来解释其产生机制。然而有意思的是，我们在大约一半的嵌合RNA的供体基因之间找到了短的同源序列，这一发现使我们提出了一种新的分子机制来解释这些嵌合RNA的形成，我们称之为“转录滑动”模型。在酵母我们，我们用实验的方法验证了短同源序列对形成嵌合RNA的必要性，有力地支持了我们这一模型。
英文摘要	The origin of new exons is a potentially important source for generating proteomic diversity. Despite recent significant progresses, many questions concerning the functional role and evolutionary importance of new exons and their host genes remain. We report a study of evolutionary and functional features of new exons and their host genes in human and mouse. We find that new exons preferentially locate in UTR especially 5’ UTR regions, implying that many new exons are involved in regulation. We also find that genes containing new exons have higher tissue specificity of gene expression and are more likely to be involved in cellular regulation and interaction with the environment. By comparing with orthologs in outgroup lineages, we were able to show that genes that gain new exons inherently evolved faster rather than gain of new exons accelerated the evolution of host genes. We report the recurrent origination of new exons in mammalian chromodomain Y like (CDYL) genes and the functional evolution associated with the new exons. The CDYL gene in the common ancestor of mammals acquired three new exons together with a new upstream promoter. Subsequently one more new exon evolved independently in mammalian lineages. In human, additional changes including start codon shift and alternative splicing occurred in CDYL gene that led to the creation of a longer peptide. The evolution of these new exons in mammals appears to be a result of positive selection as significant excess of non-synonymous mutations was observed in these exons. Functionally, the newly evolved longer peptide exhibits a weaker transcription repression activity and could attenuate the repression activity of the shorter form, suggesting that the evolution of the new exons are functionally relevant and may contribute to the complexity of the proteome. Chimeric RNAs from two or more distinct transcripts are conventionally thought to be produced by trans-splicing and have been reported in variety of organisms. We conducted a large-scale search for chimeric RNAs in the budding yeast, fly, mouse and human. Surprisingly, we identified thousands of chimeric transcripts in these organisms (except for yeast in which only five chimeric RNAs were observed), suggesting that formation of chimeric RNAs is a widespread process and can greatly contribute to the complexity of the transcriptome and proteome of organisms. However, only a small fraction (<20%) of these chimeric RNAs can be explained with the previous trans-splicing model. In contrast, we observed short homologous sequences (SHSs) at the junction sites of the source sequences for about half of the chimeric RNAs, suggesting a transcriptional slippage model. Our in vivo experiments in yeast showed that the disruption of the SHSs resulted in the disappearance of the corresponding chimeric RNAs, supporting our new model for chimeric RNAs generation.
语种	中文
公开日期	2010-11-12
内容类型	学位论文
源URL	[http://159.226.149.42:8088/handle/152453/6454]
专题	昆明动物研究所_基因起源组
推荐引用方式 GB/T 7714	李昕. 新外显子的起源与功能及短同源序列介导转录滑动产生嵌合RNA 的研究[D]. 北京. 中国科学院研究生院. 2008.

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