Involvement of the Spliceosomal U4 Small Nuclear RNA in Heterochromatic Gene Silencing at Fission Yeast Centromeres
Involvement of the Spliceosomal U4 Small Nuclear RNA in Heterochromatic Gene Silencing at Fission Yeast Centromeres
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DOI：
10.1074/jbc.m109.074393
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发表时间：
2010-02-19
影响因子：
4.8
通讯作者：
Tani, Tokio
中科院分区：
文献类型：
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作者：
Chinen, Madoka;Morita, Misato;Tani, Tokio
prp13-1 是在裂殖酵母裂殖酵母中在不允许的温度下筛选有缺陷的前 mRNA 剪接时分离出的突变体之一。我们克隆了prp13(+)基因，发现它编码参与剪接体组装的U4小核RNA(snRNA)。 prp13-1突变体产生细长的细胞，这是一种与细胞分裂周期突变体相似的表型，并且在后期纺锤体上表现出高频率的滞后染色体。该突变体对微管不稳定药物噻菌灵高度敏感，支持 prp13-1 在染色体分离方面存在缺陷。我们发现prp13-1突变导致ura4(+)基因插入着丝粒周围异染色质区域的表达，并减少异染色质蛋白Swi6p向该区域的募集，表明prp13-1中着丝粒周围异染色质的形成存在缺陷，这对于染色体的分离至关重要。粟酒裂殖酵母中的 RNA 干扰 (RNAi) 系统诱导着丝粒异染色质的形成。在prp13-1中，着丝粒非编码RNA加工成siRNA（指导异染色质形成）受到损害，并且未加工的非编码RNA积累。这些结果表明 U4 snRNA 是着丝粒处 RNAi 指导的异染色质基因沉默所必需的。关于剪接体 U4 snRNA 和 RNAi 指导的异染色质形成之间的联系，我们在着丝粒非编码 RNA 中鉴定出了 mRNA 型内含子。我们提出了一个模型，其中剪接体或亚剪接体复合物在含内含子的着丝粒非编码RNA上的组装，通过与RNA指导的RNA聚合酶复合物的相互作用，促进着丝粒处RNAi指导的异染色质的形成。
prp13-1 is one of the mutants isolated in a screen for defective pre-mRNA splicing at a nonpermissive temperature in fission yeast Schizosaccharomyces pombe. We cloned the prp13(+) gene and found that it encodes U4 small nuclear RNA (snRNA) involved in the assembly of the spliceosome. The prp13-1 mutant produced elongated cells, a phenotype similar to cell division cycle mutants, and displays a high incidence of lagging chromosomes on anaphase spindles. The mutant is hypersensitive to the microtubule-destabilizing drug thiabendazole, supporting that prp13-1 has a defect in chromosomal segregation. We found that the prp13-1 mutation resulted in expression of the ura4(+) gene inserted in the pericentromeric heterochromatin region and reduced recruitment of the heterochromatin protein Swi6p to that region, indicating defects in the formation of pericentromeric heterochromatin, which is essential for the segregation of chromosomes, in prp13-1. The formation of centromeric heterochromatin is induced by the RNA interference (RNAi) system in S. pombe. Inprp13-1, the processing of centromeric noncoding RNAs to siRNAs, which direct the heterochromatin formation, was impaired and unprocessed noncoding RNAs were accumulated. These results suggest that U4 snRNA is required for the RNAi-directed heterochromatic gene silencing at the centromeres. In relation to the linkage between the spliceosomal U4 snRNA and the RNAi-directed formation of heterochromatin, we identified a mRNA-type intron in the centromeric noncoding RNAs. We propose a model in which the assembly of the spliceosome or a sub-spliceosome complex on the intron-containing centromeric noncoding RNAs facilitates the RNAi-directed formation of heterochromatin at centromeres, through interaction with the RNA-directed RNA polymerase complex.