血清反应因子结合位点在小鼠及人基因组中保守性、多样性及进化的研究

目的：CArG元件因其为血清反应因子识别的结合位点近年来备受关注。然而迄今为止尚未见到有关CArG元件的序列特征及进化模式的研究。方法：本研究应用生物信息学方法结合遗传学方法对小鼠及人基因组中CArG元件的位置分布序列类型、多样性及保守性进行深入研究。结果：多样性研究结果显示,CArG元件的序列在小鼠及人类基因组存在大量的不同类型。但是,小鼠和人基因组中CArG元件的主要类型又存在明显差异。同源性分析结果表明人类和小鼠中的CArG元件存在两种进化历程,一部分CArG元件拥有共同的祖先,一部分是在物种分化以后突变产生的。结论：上述研究结果将为更为深入阐述SRF的调控模式奠定理论基础,同时为更清楚的阐释CArG元件序列变化对下游基因的表达影响提供理论支持。     

小鼠中血清因子结合位点位置分布及保守性的生物信息学分析

目的:血清反应因子在与心血管相关的疾病基因调控方面的作用越来越重要。血清反应因子识别的结合位点CArG元件因其重要的基因调控作用近年来随之备受关。本研究目的是揭示血清因子结合位点的位置分布与功能的关系及CArG元件内部各个位点的保守性。方法:本研究应用生物信息学方法结合遗传学方法对小鼠中CArG元件的位置分布、位点替换率及GO分类进行深入研究。结果:结果表明,71%的功能CArG元件分布在转录起始位点上游,且距离转录起始位点越近,CArG元件的数量越多。保守性分析发掘出元件内部的替换冷点、热点及替换规律。GO分类结果显示,CArG依赖性基因多为信号转导和细胞骨架蛋白。结论:上述研究结果将为准确预测候选CArG元件提供重要理论基础,同时也将为更为深入阐述SRF的调控模式奠定基础。     

小鼠中血清因子结合位点位置分布及保守性的生物信息学分析

目的：血清反应因子在与心血管相关的疾病基因调控方面的作用越来越重要。血清反应因子识别的结合位点CArG元件因其重要的基因调控作用近年来随之备受关。本研究目的是揭示血清因子结合位点的位置分布与功能的关系及CArG元件内部各个住点的保守性。方法：本研究应用生物信息学方法结合遗传学方法对小鼠中CArG元件的位置分布、位点替换率及G0分类进行深入研究。结果：结果表明,71％的功能CArG元件分布在转录起始位点上游,且距离转录起始位点越近,CArG元件的数量越多。保守性分析发掘出元件内部的替换冷点、热点及替换规律。GO分类结果显示,CArG依赖性基因多为信号转导和细胞骨架蛋白。结论：上述研究结果将为准确预测候选CArG元件提供重要理论基础,同时也将为更为深入阐述SRF的调控模式奠定基础。     

原核生物基因组肽编码sORFs分布及功能特征

近期,从非编码RNA中发现具有肽编码能力的小开放阅读框(sORFs),激发了人们对这种长期被忽略的基因组元件的研究兴趣,sORFs迅速成为当前重点研究领域.由于表达水平及丰度低、序列短等因素,对肽编码sORFs的有效研究方法及数据资源还很缺乏,现有研究仅集中在少数真核模式生物,对自然界中广泛存在的原核生物研究非常少,肽编码sORFs的发现为目前精准背景下的基因组注释提出严峻挑战.在此背景下,本文首先系统研究了80余种不同类型原核生物中长度小于100个氨基酸的肽编码sORFs分布及功能特征,并对不同长度区间sORFs的序列组成、分布及进化特征进行了对比分析.结果表明,肽编码sORFs在原核生物基因组普遍存在,随着序列长度的降低,其序列复杂度降低,行使的生物功能也相对集中.在此基础上,进一步结合当前肽编码sORFs研究现状,深入总结了肽编码sORFs研究存在的问题及挑战,为今后肽编码sORFs研究奠定了坚实理论基础.     

基于蓝藻全基因组原始数据的转座元件挖掘及组成分析

二代测序技术及全基因组多样性比较是现代生物学及信息科学研究的热点,对基因组中转座元件(Transposable element)的分析已成为基因组比较分析的重要组成部分。目前对于转座元件的种类、数量和组成的挖掘和分析一般是基于完全拼接后的全基因组序列,对在此之前的海量短片段序列后期处理及拼接仍是目前基因组研究的盲点,以转座元件为主的重复序列在拼接过程中也存在着不可避免的拼接误差或丢失,给转座元件系统的分析带来不确定。文章旨在建立一套分析流程,对铜绿微囊藻NIES 843全基因组构建的罗氏(Roche)公司454测序随机模拟原始数据集的转座元件(主要类型为插入序列:Insert sequence,IS)组成进行分析,结果表明,采用对核酸探针扫描后备选序列分成3组,并分设氨基酸检测阈值的方案分析得到的结果较为可靠,结果显示铜绿微囊藻NIES843的蓝藻转座元件占基因组比例的10.38%,归属于14个IS家族,66个IS亚家族。与之前基于完整拼接基因组数据的两套不同分析流程得到的结果相比,在丰度及家族/亚家族组成上无显著差异,在转座元件序列水平上也显示了高比例的相似性序列重叠,证实了本研究流程在基于高通量测序原始数据的转座元件分析方面具可靠性及实用性。     

原生动物基因组转座元件的研究进展

转座元件是一类广泛分布于真核生物的可移动的遗传因子, 可以引起基因重组和变异, 在物种进化及遗传改良中起着重要作用。针对近年来原生动物全基因组序列中大量发现的转座元件, 文章着重比较了转座元件在锥虫、利什曼虫、微孢子虫、变形虫和滴虫基因组序列中的存在种类、分布特征及其功能意义。原生动物转座元件以LINE 和SINE为主, 其次是DNA转座元件和LTR反转座元件, 部分转座元件在高A+T含量区富集, 预示着转座元件与基因组序列A+T含量有着紧密联系。根据不同种微孢子虫基因组之间转座元件的差异, 推测在微孢子虫基因组进化过程中, 至少经历了一次转座元件的丢失事件。最后对转座元件在原生动物寄生虫的进一步研究和应用作了展望。     

转座元件、表观遗传调控与细胞命运决定

转座元件是哺乳动物基因组内含量最多的元素。尽管转座元件的存在对基因组稳定性具有潜在的危险,但它们同时还是潜在的基因调控序列、蛋白质编码序列和染色质结构序列,并参与物种进化过程。因此,基因组中转座元件的有害性和有益性保持着谨慎的平衡,并且这种平衡主要由表观遗传修饰来调控。本文详细介绍了异染色质类型表观遗传修饰如H3K9me3和DNA甲基化在转座元件沉默中的功能;转座元件作为增强子元件富集激活型表观遗传修饰如H3K4me1和H3K27ac,以及作为转录因子结合靶点、染色质构象锚点等方式参与基因表达调控的模式;从体内胚胎发育到体外细胞命运转变,阐述了转座元件在细胞命运决定中的潜在功能及作用方式;最后,对转座元件领域研究存在的挑战及潜在解决方法提出了见解。总之,本文对转座元件与表观遗传、基因表达调控以及细胞命运决定等方面的研究及存在的问题进行了较全面的综述,旨在为相关领域的研究人员提供参考。     

人14-3-3蛋白家族的生物信息学分析

目的:分析人14-3-3蛋白家族的同源性及分子进化关系。方法:利用已公布的人基因组数据库,采用BLASTN程序检索人14-3-3蛋白家族各成员的编码基因和假基因,并利用DNAMAN软件进行序列联配,绘制其分子进化树。结果:该家族半数成员具有多个假基因序列,为返转座类型假基因。进化分析表明该家族有共同的祖先,可归为3个亚类。结论:人14-3-3蛋白家族每个成员长期进化所形成的多样性提示其功能具有独特性。     

恒河猴基因组SINEs和Alu元件的数量和分布

哺乳动物的基因组中富含各种类型的重复序列,其中的微卫星作为分子标记在相关研究中得到了广泛的应用。而重复序列中的SINEs元件,在各类群物种的分子系统发育、遗传多样性等方面的研究也得到了使用。其中,灵长类物种特有的SINEs元件、Alu元件,也在灵长类物种的进化研究中得到了成功的使用。本研究对于重要的医学模式动物恒河猴的基因组中SINEs和Alu元件进行了搜索,并进一步统计分析其分布规律、长度等信息。在恒河猴基因组的20条常染色体上共发现了Alu元件1 093 185个,在性染色体X上发现了45 215个。长度为200 bp至300 bp区间的Alu元件分布最多;Alu元件中75%的分化值至少都为10,而只有6.2%左右的元件分化值能达到至少20,这一结果表明绝大部分的Alu元件都比较年轻。本研究的统计结果为后续应用SINEs和Alu元件作为分子标记的研究提供了重要的信息。     

美洲大蠊基因组重复序列分析

重复序列是真核生物基因组的重要组成部分。一些重复序列,如自主型的逆转录转座子LINE,在昆虫的系统进化和遗传多样性研究方面得到了广泛的应用。de novo从头预测和基于同源比对预测相结合的方法被用来搜索美洲大蠊Periplaneta americana基因组,共鉴定出大约占全基因组62%的重复序列。研究发现,散在重复序列中,DNA转座子占美洲大蠊基因组的16.18%;逆转座元件中LINE最多,占基因组的13.64%,SINE和LTR分别占基因组的3.52%和1.32%。LINEs中的Bov Bs亚家族在所有转座子亚家族中比例最高(约6.73%)。美洲大蠊与德国小蠊Blattella germanica相比,除LTR外,其他类型的转座子占基因组的比例均高于德国小蠊。通过分析逆转录转座子反转录酶完整度、氨基酸序列相似度及遗传距离,从美洲大蠊基因组中鉴定出一类BovBs:RTE-1_PAm。BovBs的反转录酶氨基酸序列的系统树表明,美洲大蠊与内华达古白蚁Zootermopsis nevadensis的进化关系比与其同属蜚蠊科Blattidae的德国小蠊的关系更近。昆虫中BovBs的进化关系与传统核基因进化关系的不同,表明转座子的进化相对宿主基因的进化具有一定的独立性。     

Substitution pattern of the CArG element in human and mouse genomes

cis-Elements CArG bound by serum response factor (SRF) are presently being intensively studied, but little is known about the substitution pattern of functional CArG elements. Here, we have performed the first evolutionary analysis of CArGome in the human and mouse genome through bioinformatic methods and statistical tests. We calculated the substitution rate at each site of the functional CArG elements. The results showed that the core sites of the functional CArG elements evolved faster than did the background DNA, indicating that these sites were likely to evolve under positive selection. Moreover, a strong TATA "motif" was evident in the core region within the functional CArG elements in both human and mouse promoters. This motif could probably be a major contribution to the formation of the spatial structure, which was important for CArG-SRF recognition. Thus, the study further revealed the sequence character and substitution pattern of CArG elements and provided useful information for the study of the SRF-binding efficiencies of CArG promoters in functional assays.     

The population biology of transposable elements

A transposable element can be defined as a DNA sequence capable of moving to new sites in the genome. Such DNA sequences have been described in a wide range of organisms. The evolutionary processes affecting transposable elements can thus be divided into two categories: changes in sequence and changes in genomic location. As with other types of evolutionary change, the nature of the evolutionary process will be reflected in the extent and type of genetic variation existing in wild populations. Quantitative models of the evolution of transposable element sequences and positions will be outlined, and related to relevant data. The extent to which models designed to describe obvious transposable elements such as the mobile sequences of Drosophila are also applicable to interspersed repetitive DNAs from other species will be discussed.     

Identification of a conserved sequence in the non-coding regions of many human genes.

We have analyzed a sequence of approximately 70 base pairs (bp) that shows a high degree of similarity to sequences present in the non-coding regions of a number of human and other mammalian genes. The sequence was discovered in a fragment of human genomic DNA adjacent to an integrated hepatitis B virus genome in cells derived from human hepatocellular carcinoma tissue. When one of the viral flanking sequences was compared to nucleotide sequences in GenBank, more than thirty human genes were identified that contained a similar sequence in their non-coding regions. The sequence element was usually found once or twice in a gene, either in an intron or in the 5' or 3' flanking regions. It did not share any similarities with known short interspersed nucleotide elements (SINEs) or presently known gene regulatory elements. This element was highly conserved at the same position within the corresponding human and mouse genes for myoglobin and N-myc, indicating evolutionary conservation and possible functional importance. Preliminary DNase I footprinting data suggested that the element or its adjacent sequences may bind nuclear factors to generate specific DNase I hypersensitive sites. The size, structure, and evolutionary conservation of this sequence indicates that it is distinct from other types of short interspersed repetitive elements. It is possible that the element may have a cis-acting functional role in the genome.     

Phylogenetic Analysis of a Retroposon Family in African Great Apes

The SINE-R retroposon family has been identified by its relationship with the long terminal repeats (LTRs) of human endogenous retrovirus class K (HERV-K) as a mobile element that has evolved recently in the human genome. Here we examined the recent evolutionary history of this class of elements by a PCR approach to genomic DNA from the African great apes and by phylogenetic analysis including comparison with the HERV K10 parent sequence. With primers derived from a cDNA sequence from human brain, we identified 27 sequences from the chimpanzee and 16 from the gorilla. Phylogenetic comparisons with previously recognized sequences from the human and from the orangutan and gibbon revealed wide overlap of elements across species, suggesting multiple origins in the course of hominoid evolution. Two human elements SINE-R.C2 and HS307 were the furthest removed from the HERV-K10 sequence but these two elements were closely related to three elements from the chimpanzee and four elements from the gorilla. This group of elements (our clusters 14 and 15) appears to have transposed late in hominoid evolution. One element (Ch-M16) showed 99.1% sequence identity with the SINE-R.C2 element, which is human-specific. Thus the SINE-R family appears to have continued to be active in transposition throughout the course of primate evolution. Received: 12 March 1999 / Accepted: 25 May 1999     

Natural and synthetic DNA elements with the CArG motif differ in expression and protein-binding properties.

DNA elements with the CC(A/T)6GG, or CArG, motif occur in promoters that are under different regulatory controls. CArG elements from the skeletal actin, c-fos, and myogenin genes were tested for their abilities to confer tissue-specific expression on reporter genes when the individual elements were situated immediately upstream from a TATA element. The c-fos CArG element, also referred to as the serum response element (SRE), conferred basal, constitutive expression on the test promoter. The CArG motif from the myogenin gene was inactive. The skeletal actin CArG motif functioned as a muscle regulatory element (MRE) in that basal expression was detected only in muscle cultures. Muscle-specific expression from the 28-bp MRE and the 2.3-kb skeletal actin promoter was trans repressed by the Fos and Jun proteins. The expression and factor-binding properties of a series of synthetic CArG elements were analyzed. Muscle-specific expression was conferred by perfect 28-bp palindromes on the left and right halves of the skeletal actin MRE. Chimeric elements of the skeletal actin MRE and the c-fos SRE differed in their expression properties. Muscle-specific expression was observed when the left half of the MRE was fused to the right half of the SRE. Constitutive expression was conferred by a chimera with the right half of the MRE fused to the left half of the SRE and by chimeras which exchanged the central CC(A/T)6GG sequences. At least three distinct proteins specifically bound to these CArG elements. The natural and synthetic CArG elements differed in their affinities for these proteins; however, muscle-specific expression could not be attributed to differences in the binding of a single protein. Furthermore, the MRE did not bind MyoD or the myogenin-E12 heterodimer, indicating that muscle-specific expression from this element does not involve a direct interaction with these helix-loop-helix proteins. These data demonstrate that the conserved CArG motifs form the core of a family of functionally different DNA regulatory elements that may contribute to the tissue-specific expression properties of their cognate promoters.