The Prp19-associated complex is required for specifying interactions of U5 and U6 with pre-mRNA during spliceosome activation
The Prp19-associated complex is required for specifying interactions of U5 and U6 with pre-mRNA during spliceosome activation
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DOI：
10.1074/jbc.m505060200
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发表时间：
2005-09-02
影响因子：
4.8
通讯作者：
Cheng, SC
中科院分区：
文献类型：
--
作者：
Chan, SP;Cheng, SC
剪接体的激活涉及剪接体中的主要结构变化，包括释放U1和U4小的核核糖核蛋白颗粒以及添加大的蛋白质复合物Prp 19相关复合物。我们以前表明，Prp 19相关的复合物是U4释放后U 5和U6与剪接体稳定结合所必需的。Prp 19相关复合物结合后剪接体内的变化包括U6/5'剪接位点相互作用的重塑和Lsm蛋白的不稳定，以允许U6与内含子序列进一步相互作用。在此，我们进一步分析了U 5和U6与前mRNA在剪接体组装的各个阶段的相互作用，从三小核核糖核蛋白复合物的初始结合到激活的剪接体，以揭示相互作用的逐步变化。我们证明，U 5和U6与前mRNA的动态方式跨越了一个大区域的U6和5'外显子序列的剪接体激活之前的相互作用。在剪接体激活过程中，相互作用被锁定在小区域，Prp 19相关复合物是确定U 5和U6与5'剪接位点相互作用的特异性所必需的，以在U4解离后稳定它们与剪接体的结合。
Activation of the spliceosome involves a major structural change in the spliceosome, including release of U1 and U4 small nuclear ribonucleoprotein particles and the addition of a large protein complex, the Prp19-associated complex. We previously showed that the Prp19-associated complex is required for stable association of U5 and U6 with the spliceosome after U4 is released. Changes within the spliceosome upon binding of the Prp19-associated complex include remodeling of the U6/5' splice site interaction and destabilization of Lsm proteins to allow further interaction of U6 with the intron sequence. Here, we further analyzed interactions of U5 and U6 with pre-mRNA at various stages of spliceosome assembly from initial binding of tri- small nuclear ribonucleoprotein complex to the activated spliceosome to reveal stepwise changes of interactions. We demonstrate that both U5 and U6 interacted with pre-mRNA in dynamic manners spanning over a large region of U6 and the 5' exon sequences prior to the activation of the spliceosome. During spliceosome activation, interactions were locked down to small regions, and the Prp19-associated complex was required for defining the specificity of interaction of U5 and U6 with the 5' splice site to stabilize their association with the spliceosome after U4 is dissociated.