Splicing-dependent and -independent modes of assembly for intron-encoded box C/D snoRNPs in mammalian cells
Splicing-dependent and -independent modes of assembly for intron-encoded box C/D snoRNPs in mammalian cells
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DOI：
10.1016/s1097-2765(03)00267-3
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发表时间：
2003-07-01
期刊：
影响因子：
16
通讯作者：
Steitz, JA
中科院分区：
文献类型：
--
作者：
Hirose, T;Shu, MD;Steitz, JA
在哺乳动物细胞中，所有指导rRNA修饰的小核仁rna （snoRNAs）都编码在宿主基因的内含子内。宿主内含子3'剪接位点上游约70 nts的最佳位置对于体内box C/D snoRNAs的高效表达至关重要，这表明它与剪接有协同作用。在这里，我们使用体外剪接-snoRNA加工系统来证明盒C/D snoRNP蛋白的组装是受snoRNA位置影响的步骤，并且主动剪接对于snoRNP组装至关重要。剪接阻断实验进一步揭示了snoRNP蛋白在剪接体C1复合体阶段特异性结合。相比之下，在体外具有稳定外茎的snoRNAs上可以发生不依赖剪接的snoRNP组装。体内分析证实，一个稳定的系统可以弥补那些远离宿主内含子3'剪接位点的少数盒状C/D snorna的不寻常位置。
In mammalian cells, all small nucleolar RNAs (snoRNAs) that guide rRNA modification are encoded within the introns of host genes. An optimal position about 70 nts upstream of the 3' splice site of the host intron is critical for efficient expression of box C/D snoRNAs in vivo, suggesting synergy with splicing. Here, we have used a coupled in vitro splicing-snoRNA processing system to demonstrate that assembly of box C/D snoRNP proteins is the step affected by snoRNA location, and that active splicing is essential for snoRNP assembly. Splicing blockage experiments further reveal that snoRNP proteins bind specifically at the spliceosomal C1 complex stage. In contrast, splicing-independent snoRNP assembly can occur in vitro on snoRNAs that possess stable external stems. In vivo analyses confirm that a stable stem can compensate for the unusual position of those few box C/D snoRNAs located far from the 3' splice site of their host intron.