Discovery and analysis of evolutionarily conserved intronic splicing regulatory elements.
Discovery and analysis of evolutionarily conserved intronic splicing regulatory elements.
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DOI：
10.1371/journal.pgen.0030085
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发表时间：
2007-05-25
期刊：
影响因子：
4.5
通讯作者：
Liang, Tiffany Y.
中科院分区：
文献类型：
--
作者：
Yeo, Gene W.;Van Nostrand, Eric L.;Liang, Tiffany Y.
了解调节结构性和选择性前mRNA剪接的顺式调控元件是生物学和医学的基础。在这里，我们采用了全基因组比较基因组学的方法，利用现有的哺乳动物基因组来确定保守的内含子剪接调控元件(ISRES)。我们的方法产生了314个ISRES，在相互竞争的剪接位点之间插入~70个ISRES表明，84%的ISRES改变了人类细胞的5‘和94%的剪接位点选择。与我们的实验一致，ISRES与已知剪接调控元件的比较表明，40%-45%的ISRES可能具有外显子剪接沉默的双重作用。支持ISRES在选择性剪接中的作用，我们发现30%-50%的ISRES在选择性剪接(AS)外显子附近富含，并且几乎包括所有已知的组织特异性选择性剪接因子的结合位点。此外，我们观察到含有ISRE近端外显子的基因对ISRE特有的组织表达和分子功能具有偏见。最后，我们发现，对于Nova1、神经元PTB、hnRNP C和FOX1，剪接因子中另一个保守外显子附近最频繁出现的ISRE与其自身已知的RNA结合位点非常相似，这表明ISRE密度和剪接因子自动调节的倾向将RNA结合位点与剪接因子关联起来是一种新的应用。我们的结果表明，ISRES是理解一般性和组织特异性AS调控以及由这些AS调控的生物学途径和功能的关键构件。在RNA剪接过程中，前mRNA中的序列(内含子)被切除并丢弃，其余序列(外显子)连接起来形成成熟的RNA。剪接不仅受基本剪接机制与位于外显子-内含子边界的剪接位点的结合调控，而且受各种其他剪接因子的共同影响，这些剪接因子结合位于外显子和侧翼内含子中的大量序列元件。选择性剪接的实例，其中剪接位点(S)的使用不完整或在组织、细胞类型或谱系之间不同，可以通过序列元件与组织、细胞类型和阶段特异性剪接因子的相互作用而产生。为了更好地了解结构性和选择性的前mRNA剪接，作者描述了一种比较基因组学方法，利用现有的哺乳动物基因组，系统地确定位于外显子近端内含子的剪接调控元件。四分之一的元素进行了实验测试，其中大多数都改变了人类细胞的剪接。作者还表明，内含子元件接近组织特异性的替代外显子，更有可能位于内含子的特定位置，提示潜在的调节功能。这些元件也经常在组织特异性基因中发现，这表明这些基因的表达和选择性剪接之间存在耦合。最后，作者提出了一种利用这些元件来识别几种剪接因子的结合位点的策略。
Knowledge of the functional cis-regulatory elements that regulate constitutive and alternative pre-mRNA splicing is fundamental for biology and medicine. Here we undertook a genome-wide comparative genomics approach using available mammalian genomes to identify conserved intronic splicing regulatory elements (ISREs). Our approach yielded 314 ISREs, and insertions of ~70 ISREs between competing splice sites demonstrated that 84% of ISREs altered 5′ and 94% altered 3′ splice site choice in human cells. Consistent with our experiments, comparisons of ISREs to known splicing regulatory elements revealed that 40%–45% of ISREs might have dual roles as exonic splicing silencers. Supporting a role for ISREs in alternative splicing, we found that 30%–50% of ISREs were enriched near alternatively spliced (AS) exons, and included almost all known binding sites of tissue-specific alternative splicing factors. Further, we observed that genes harboring ISRE-proximal exons have biases for tissue expression and molecular functions that are ISRE-specific. Finally, we discovered that for Nova1, neuronal PTB, hnRNP C, and FOX1, the most frequently occurring ISRE proximal to an alternative conserved exon in the splicing factor strongly resembled its own known RNA binding site, suggesting a novel application of ISRE density and the propensity for splicing factors to auto-regulate to associate RNA binding sites to splicing factors. Our results demonstrate that ISREs are crucial building blocks in understanding general and tissue-specific AS regulation and the biological pathways and functions regulated by these AS events. During RNA splicing, sequences (introns) in a pre-mRNA are excised and discarded, and the remaining sequences (exons) are joined to form the mature RNA. Splicing is regulated not only by the binding of the basic splicing machinery to splice sites located at the exon–intron boundaries, but also by the combined effects of various other splicing factors that bind to a multitude of sequence elements located both in the exons as well as the flanking introns. Instances of alternative splicing, where usage of splice site(s) is incomplete or different between tissues, cell types, or lineages, can be created by the interaction of sequence elements and tissue, cell type, and stage-specific splicing factors. To better understand constitutive and alternative pre-mRNA splicing, the authors describe a comparative genomics approach, using available mammalian genomes, to systematically identify splicing regulatory elements located in the introns proximal to exons. A quarter of the elements were tested experimentally, and most of them altered splicing in human cells. The authors also showed that that the intronic elements are close to tissue-specific alternative exons and are more likely to be located in specific positions in the introns, suggestive of potential regulatory function. These elements are also frequently found in tissue-specific genes, suggesting a coupling between expression and alternative splicing of these genes. Finally, the authors propose a strategy using the elements to identify the binding sites of several splicing factors.
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DOI：
10.1073/pnas.94.23.12343
发表时间：
1997-11-11
影响因子：
11.1
作者：
Hedjran, F;Yeakley, JM;Rosenfeld, MG
通讯作者：
Rosenfeld, MG
影响因子：
10.5
作者：
HUH, GS;HYNES, RO
通讯作者：
HYNES, RO
影响因子：
64.8
作者：
Kellis, M;Patterson, N;Lander, ES
通讯作者：
Lander, ES
影响因子：
11.4
作者：
Dredge, BK;Stefani, G;Darnell, RB
通讯作者：
Darnell, RB
影响因子：
11.4
作者：
Jin, Y;Suzuki, H;Inoue, K
通讯作者：
Inoue, K