Plass Mireya, Eyras Eduardo
Research Unit of Biomedical Informatics, IMIM - Pompeu Fabra University, E08003, Barcelona, Spain.
BMC Evol Biol. 2006 Jun 22;6:50. doi: 10.1186/1471-2148-6-50.
Alternatively spliced exons play an important role in the diversification of gene function in most metazoans and are highly regulated by conserved motifs in exons and introns. Two contradicting properties have been associated to evolutionary conserved alternative exons: higher sequence conservation and higher rate of non-synonymous substitutions, relative to constitutive exons. In order to clarify this issue, we have performed an analysis of the evolution of alternative and constitutive exons, using a large set of protein coding exons conserved between human and mouse and taking into account the conservation of the transcript exonic structure. Further, we have also defined a measure of the variation of the arrangement of exonic splicing enhancers (ESE-conservation score) to study the evolution of splicing regulatory sequences. We have used this measure to correlate the changes in the arrangement of ESEs with the divergence of exon and intron sequences.
We find evidence for a relation between the lack of conservation of the exonic structure and the weakening of the sequence evolutionary constraints in alternative and constitutive exons. Exons in transcripts with non-conserved exonic structures have higher synonymous (dS) and non-synonymous (dN) substitution rates than exons in conserved structures. Moreover, alternative exons in transcripts with non-conserved exonic structure are the least constrained in sequence evolution, and at high EST-inclusion levels they are found to be very similar to constitutive exons, whereas alternative exons in transcripts with conserved exonic structure have a dS significantly lower than average at all EST-inclusion levels. We also find higher conservation in the arrangement of ESEs in constitutive exons compared to alternative ones. Additionally, the sequence conservation at flanking introns remains constant for constitutive exons at all ESE-conservation values, but increases for alternative exons at high ESE-conservation values.
We conclude that most of the differences in dN observed between alternative and constitutive exons can be explained by the conservation of the transcript exonic structure. Low dS values are more characteristic of alternative exons with conserved exonic structure, but not of those with non-conserved exonic structure. Additionally, constitutive exons are characterized by a higher conservation in the arrangement of ESEs, and alternative exons with an ESE-conservation similar to that of constitutive exons are characterized by a conservation of the flanking intron sequences higher than average, indicating the presence of more intronic regulatory signals.
可变剪接外显子在大多数后生动物的基因功能多样化中发挥着重要作用，并受到外显子和内含子中保守基序的高度调控。相对于组成型外显子，进化保守的可变外显子具有两种相互矛盾的特性：更高的序列保守性和更高的非同义替换率。为了阐明这个问题，我们利用一组在人类和小鼠之间保守的蛋白质编码外显子，并考虑转录本外显子结构的保守性，对外显子和组成型外显子的进化进行了分析。此外，我们还定义了一种外显子剪接增强子排列变化的度量（ESE保守分数），以研究剪接调控序列的进化。我们使用这种度量来关联ESE排列的变化与外显子和内含子序列的分歧。
我们发现证据表明，外显子结构缺乏保守性与可变外显子和组成型外显子中序列进化约束的减弱之间存在关联。外显子结构不保守的转录本中的外显子比保守结构中的外显子具有更高的同义（dS）和非同义（dN）替换率。此外，外显子结构不保守的转录本中的可变外显子在序列进化中受到的约束最小，在高EST包含水平下，它们与组成型外显子非常相似，而外显子结构保守的转录本中的可变外显子在所有EST包含水平下的dS都显著低于平均水平。我们还发现，与可变外显子相比，组成型外显子中ESE的排列具有更高的保守性。此外，对于组成型外显子，侧翼内含子的序列保守性在所有ESE保守值下保持不变，但对于高ESE保守值下的可变外显子，其序列保守性会增加。
我们得出结论，可变外显子和组成型外显子之间观察到的大部分dN差异可以通过转录本外显子结构的保守性来解释。低dS值更具外显子结构保守的可变外显子的特征，但不具有外显子结构不保守的可变外显子的特征。此外，组成型外显子的特征是ESE排列具有更高保守性，而ESE保守性与组成型外显子相似的可变外显子的特征是侧翼内含子序列的保守性高于平均水平，这表明存在更多的内含子调控信号。
