Interconnections Between RNA-Processing Pathways Revealed by a Sequencing-Based Genetic Screen for Pre-mRNA Splicing Mutants in Fission Yeast.
Interconnections Between RNA-Processing Pathways Revealed by a Sequencing-Based Genetic Screen for Pre-mRNA Splicing Mutants in Fission Yeast.
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DOI：
10.1534/g3.116.027508
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发表时间：
2016-06-01
期刊：
影响因子：
--
通讯作者：
Pleiss JA
中科院分区：
文献类型：
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作者：
Larson A;Fair BJ;Pleiss JA
前mrna剪接是真核生物基因表达的重要组成部分，从单细胞酵母到人类都高度保守。在这里，我们提出了一种基于测序的反遗传筛选的开发和实施，旨在鉴定影响分裂酵母裂糖酵母（Schizosaccharomyces pombe）前mrna剪接的非必需基因，裂糖酵母是一种具有高等真核生物剪接许多复杂特征的生物。使用定制设计的条形码方案，我们同时查询了约3000个突变株，以了解它们对两种内源性pre- mrna剪接效率的影响。共鉴定出61个非必需基因，其缺失导致前mrna剪接缺陷；其中富含编码剪接体已知或预测组分的因子。在这里确定的候选基因中包括在其他rna加工途径中具有良好特征的基因，包括异色沉默和3′末端加工。剪接敏感微阵列证实了许多这些因素的广泛剪接缺陷，揭示了这些途径之间新的功能联系。
Pre-mRNA splicing is an essential component of eukaryotic gene expression and is highly conserved from unicellular yeasts to humans. Here, we present the development and implementation of a sequencing-based reverse genetic screen designed to identify nonessential genes that impact pre-mRNA splicing in the fission yeast Schizosaccharomyces pombe, an organism that shares many of the complex features of splicing in higher eukaryotes. Using a custom-designed barcoding scheme, we simultaneously queried ∼3000 mutant strains for their impact on the splicing efficiency of two endogenous pre-mRNAs. A total of 61 nonessential genes were identified whose deletions resulted in defects in pre-mRNA splicing; enriched among these were factors encoding known or predicted components of the spliceosome. Included among the candidates identified here are genes with well-characterized roles in other RNA-processing pathways, including heterochromatic silencing and 3ʹ end processing. Splicing-sensitive microarrays confirm broad splicing defects for many of these factors, revealing novel functional connections between these pathways.