In vivo commitment to yeast cotranscriptiona splicing is sensitive to transcription elongation mutants
In vivo commitment to yeast cotranscriptiona splicing is sensitive to transcription elongation mutants
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DOI：
10.1101/gad.1434706
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发表时间：
2006-08-01
影响因子：
10.5
通讯作者：
Rosbash, Michael
中科院分区：
文献类型：
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作者：
Lacadie, Scott A.;Tardiff, Daniel F.;Rosbash, Michael
最近研究表明，芽殖酵母酿酒酵母中的剪接体组装发生在转录位点。然而，共转录剪接以及转录和剪接之间的耦合的证据仍然缺乏。使用修改以前发表的染色质免疫沉淀（ChIP）测定，我们表明，共转录剪接发生类似于1 kb后转录的3 '剪接位点（3 ' SS）。该途径还保护大多数含内含子的新生转录物免受内含子锤头状核酶切割的影响。这表明高百分比的内含子被共转录识别。这一观察结果使我们筛选一个小的缺失文库，用于使剪接报告基因对核酶切割敏感的菌株。Delta mud2菌株的表征表明，早期剪接因子Mud2p与U1 snRNP一起作用，在新生前mRNA上形成跨内含子桥接复合物。该复合物有助于保护转录物免受核酶介导的破坏，并表明剪接体组装过程早期的内含子定义事件。转录延伸突变株Delta dst 1和Delta paf 1显示不同的共转录剪接表型，表明不同的转录途径差异影响新生内含子定义的效率。
Spliceosome assembly in the budding yeast Saccharomyces cerevisiae was recently shown to occur at the site of transcription. However, evidence for cotranscriptional splicing as well as for coupling between transcription and splicing is still lacking. Using modifications of a previously published chromatin immunoprecipitation (ChIP) assay, we show that cotranscriptional splicing occurs similar to 1 kb after transcription of the 3 ' splice site (3 ' SS). This pathway furthermore protects most intron-containing nascent transcripts from the effects of cleavage by an intronic hammerhead ribozyme. This suggests that a high percentage of introns are recognized cotranscriptionally. This observation led us to screen a small deletion library for strains that sensitize a splicing reporter to ribozyme cleavage. Characterization of the Delta mud2 strain indicates that the early splicing factor Mud2p functions with U1 snRNP to form a cross-intron bridging complex on nascent pre-mRNA. The complex helps protect the transcript from ribozyme-mediated destruction and suggests an intron-definition event early in the spliceosome assembly process. The transcription elongation mutant strains Delta dst1 and Delta paf1 show different cotranscriptional splicing phenotypes, suggesting that different transcription pathways differentially impact the efficiency of nascent intron definition.