A genome-wide analysis indicates that yeast pre-mRNA splicing is predominantly post-transcriptional

Recent ChIP experiments indicate that spliceosome assembly and splicing can occur cotranscriptionally in S. cerevisiae. However, only a few genes have been examined, and all have long second exons. To extend these studies, we analyzed intron-containing genes with different second exon lengths, by ChIP as well as by whole-genome tiling arrays (ChIP-CHIP). The data indicate that U1 snRNP recruitment is independent of exon length. Recursive splicing constructs, which uncouple U1 recruitment from transcription, suggest that cotranscriptional U1 recruitment contributes to optimal splicing efficiency. In contrast, U2 snRNP recruitment as well as cotranscriptional splicing is deficient on short second exon-genes. We estimate that approximately 90% of endogenous yeast splicing is post-transcriptional, consistent with an analysis of post-transcriptional snRNP-associated pre-mRNA. Keywords: ChIP-CHIP ChIP-CHIP was performed on tagged U1, U2, and U5 snRNPs to address spliceosome assembly on intron-containing genes in S. cerevisiae. Data expressed as log2 (immunoprecipitation/input) ratios.

近期的染色质免疫共沉淀（Chromatin Immunoprecipitation, ChIP）实验表明，酿酒酵母（Saccharomyces cerevisiae, S. cerevisiae）的剪接体组装与剪接过程可在共转录阶段完成。然而目前仅对少数基因开展了相关研究，且所有被检测基因的第二外显子均较长。为拓展此类研究范畴，本研究采用染色质免疫共沉淀（ChIP）与全基因组平铺芯片（whole-genome tiling arrays, ChIP-CHIP）技术，对第二外显子长度各异的含内含子基因展开了分析。数据显示，U1小核核糖核蛋白颗粒（U1 small nuclear ribonucleoprotein particle, U1 snRNP）的招募与外显子长度无关。采用可将U1招募与转录过程解偶联的递归剪接构建体开展实验，结果提示共转录阶段的U1招募有助于实现最优剪接效率。与之相反，在第二外显子较短的基因中，U2小核核糖核蛋白颗粒（U2 snRNP）的招募以及共转录剪接过程均存在缺陷。本研究估算，酿酒酵母中约90%的内源剪接事件均为转录后发生，这一结论与针对转录后结合有snRNP的前体mRNA的分析结果相符。关键词：ChIP-CHIP
本研究对带有标签的U1、U2及U5 snRNP开展染色质免疫共沉淀实验，以探究酿酒酵母中含内含子基因的剪接体组装过程。实验数据以log₂（免疫沉淀/输入）比值形式呈现。