Jády Beáta E, Darzacq Xavier, Tucker Karen E, Matera A Gregory, Bertrand Edouard, Kiss Tamás
Laboratoire de Biologie Moléculaire Eucaryote du CNRS, Université Paul Sabatier, 118 route de Narbonne, 31062 Toulouse Cedex, France.
EMBO J. 2003 Apr 15;22(8):1878-88. doi: 10.1093/emboj/cdg187.
Biogenesis of functional spliceosomal small nuclear RNAs (snRNAs) includes the post-transcriptional covalent modification of numerous internal nucleotides. We have recently demonstrated that synthesis of 2'-O-methylated nucleotides and pseudouridines in the RNA polymerase II-synthesized Sm snRNAs is directed by sequence-specific guide RNAs. Here, we provide evidence supporting the notion that modification of Sm snRNAs occurs in nucleoplasmic Cajal bodies (CBs), where modification guide RNAs accumulate. We show that short fragments of Sm snRNAs are correctly and efficiently modified when targeted to CBs, but not when these same fragments are targeted to the nucleolus. We also demonstrate that internal modification of the U2 snRNA occurs exclusively after nuclear import of the newly assembled Sm snRNP from the cytoplasm. Finally, we show that p80 coilin, the CB marker protein, is not required for snRNA modification. In coilin knockout cells, Sm snRNAs and their modification guide RNAs colocalize in residual CBs, which do not stockpile fibrillarin and fail to recruit the U3 small nucleolar RNA.
功能性剪接体小核RNA（snRNA）的生物合成包括许多内部核苷酸的转录后共价修饰。我们最近证明，RNA聚合酶II合成的Sm snRNA中2'-O-甲基化核苷酸和假尿苷的合成是由序列特异性引导RNA指导的。在这里，我们提供证据支持Sm snRNA的修饰发生在核质卡哈尔体（CB）中的观点，修饰引导RNA在那里积累。我们表明，Sm snRNA的短片段靶向CB时能正确且高效地被修饰，但靶向核仁时则不然。我们还证明，U2 snRNA的内部修饰仅在新组装的Sm snRNP从细胞质进行核输入后发生。最后，我们表明，CB标记蛋白p80卷曲螺旋蛋白对于snRNA修饰并非必需。在卷曲螺旋蛋白基因敲除细胞中，Sm snRNA及其修饰引导RNA共定位于残留的CB中，这些CB不储存纤维蛋白原，也无法招募U3小核仁RNA。
