Koren Eli, Lev-Maor Galit, Ast Gil
Department of Human Molecular Genetics, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel.
PLoS Comput Biol. 2007 May;3(5):e95. doi: 10.1371/journal.pcbi.0030095.
Alternative 3' and 5' splice site (ss) events constitute a significant part of all alternative splicing events. These events were also found to be related to several aberrant splicing diseases. However, only few of the characteristics that distinguish these events from alternative cassette exons are known currently. In this study, we compared the characteristics of constitutive exons, alternative cassette exons, and alternative 3'ss and 5'ss exons. The results revealed that alternative 3'ss and 5'ss exons are an intermediate state between constitutive and alternative cassette exons, where the constitutive side resembles constitutive exons, and the alternative side resembles alternative cassette exons. The results also show that alternative 3'ss and 5'ss exons exhibit low levels of symmetry (frame-preserving), similar to constitutive exons, whereas the sequence between the two alternative splice sites shows high symmetry levels, similar to alternative cassette exons. In addition, flanking intronic conservation analysis revealed that exons whose alternative splice sites are at least nine nucleotides apart show a high conservation level, indicating intronic participation in the regulation of their splicing, whereas exons whose alternative splice sites are fewer than nine nucleotides apart show a low conservation level. Further examination of these exons, spanning seven vertebrate species, suggests an evolutionary model in which the alternative state is a derivative of an ancestral constitutive exon, where a mutation inside the exon or along the flanking intron resulted in the creation of a new splice site that competes with the original one, leading to alternative splice site selection. This model was validated experimentally on four exons, showing that they indeed originated from constitutive exons that acquired a new competing splice site during evolution.
可变3'和5'剪接位点（ss）事件构成了所有可变剪接事件的重要组成部分。这些事件还被发现与几种异常剪接疾病有关。然而，目前仅知道少数几个将这些事件与可变盒式外显子区分开来的特征。在本研究中，我们比较了组成型外显子、可变盒式外显子以及可变3'ss和5'ss外显子的特征。结果显示，可变3'ss和5'ss外显子处于组成型外显子和可变盒式外显子之间的中间状态，其组成型一侧类似于组成型外显子，可变一侧类似于可变盒式外显子。结果还表明，可变3'ss和5'ss外显子表现出与组成型外显子相似的低水平对称性（读框保留），而两个可变剪接位点之间的序列则表现出与可变盒式外显子相似的高水平对称性。此外，侧翼内含子保守性分析表明，其可变剪接位点相距至少九个核苷酸的外显子具有较高的保守水平，表明内含子参与了其剪接调控，而可变剪接位点相距少于九个核苷酸的外显子则具有较低的保守水平。对跨越七个脊椎动物物种的这些外显子的进一步研究提出了一种进化模型，其中可变状态是祖先组成型外显子的衍生物，外显子内部或侧翼内含子中的突变导致产生了一个与原始剪接位点竞争的新剪接位点，从而导致可变剪接位点的选择。该模型在四个外显子上通过实验得到了验证，表明它们确实起源于在进化过程中获得了一个新的竞争剪接位点的组成型外显子。
