Erkelenz Steffen, Theiss Stephan, Otte Marianne, Widera Marek, Peter Jan Otto, Schaal Heiner
Institute for Virology, Heinrich-Heine-University Duesseldorf, Duesseldorf, Germany.
Institute of Clinical Neuroscience and Medical Psychology, Heinrich-Heine-University Duesseldorf, Duesseldorf, Germany.
Nucleic Acids Res. 2014;42(16):10681-97. doi: 10.1093/nar/gku736. Epub 2014 Aug 21.
Effective splice site selection is critically controlled by flanking splicing regulatory elements (SREs) that can enhance or repress splice site use. Although several computational algorithms currently identify a multitude of potential SRE motifs, their predictive power with respect to mutation effects is limited. Following a RESCUE-type approach, we defined a hexamer-based 'HEXplorer score' as average Z-score of all six hexamers overlapping with a given nucleotide in an arbitrary genomic sequence. Plotted along genomic regions, HEXplorer score profiles varied slowly in the vicinity of splice sites. They reflected the respective splice enhancing and silencing properties of splice site neighborhoods beyond the identification of single dedicated SRE motifs. In particular, HEXplorer score differences between mutant and reference sequences faithfully represented exonic mutation effects on splice site usage. Using the HIV-1 pre-mRNA as a model system highly dependent on SREs, we found an excellent correlation in 29 mutations between splicing activity and HEXplorer score. We successfully predicted and confirmed five novel SREs and optimized mutations inactivating a known silencer. The HEXplorer score allowed landscaping of splicing regulatory regions, provided a quantitative measure of mutation effects on splice enhancing and silencing properties and permitted calculation of the mutationally most effective nucleotide.
有效的剪接位点选择受到侧翼剪接调控元件（SREs）的严格控制，这些元件可以增强或抑制剪接位点的使用。虽然目前有几种计算算法可以识别大量潜在的SRE基序，但它们对突变效应的预测能力有限。采用一种类似RESCUE的方法，我们将基于六聚体的“HEXplorer评分”定义为与任意基因组序列中给定核苷酸重叠的所有六个六聚体的平均Z评分。沿着基因组区域绘制，HEXplorer评分图谱在剪接位点附近变化缓慢。它们反映了剪接位点邻域各自的剪接增强和沉默特性，而不仅仅是识别单个特定的SRE基序。特别是，突变序列和参考序列之间的HEXplorer评分差异忠实地反映了外显子突变对剪接位点使用的影响。以高度依赖SREs的HIV-1前体mRNA为模型系统，我们发现29个突变的剪接活性与HEXplorer评分之间具有极好的相关性。我们成功预测并证实了五个新的SREs，并优化了使已知沉默子失活的突变。HEXplorer评分能够描绘剪接调控区域，提供了一种对剪接增强和沉默特性的突变效应进行定量测量的方法，并允许计算突变最有效的核苷酸。
