Fachbereich Biologie/Chemie/Pharmazie, Abteilung Strukturbiochemie, Freie Universität Berlin, D-14195 Berlin, Germany;
Genes Dev. 2011 Aug 1;25(15):1601-12. doi: 10.1101/gad.635911. Epub 2011 Jul 15.
Little is known about how particle-specific proteins are assembled on spliceosomal small nuclear ribonucleoproteins (snRNPs). Brr2p is a U5 snRNP-specific RNA helicase required for spliceosome catalytic activation and disassembly. In yeast, the Aar2 protein is part of a cytoplasmic precursor U5 snRNP that lacks Brr2p and is replaced by Brr2p in the nucleus. Here we show that Aar2p and Brr2p bind to different domains in the C-terminal region of Prp8p; Aar2p interacts with the RNaseH domain, whereas Brr2p interacts with the Jab1/MPN domain. These domains are connected by a long, flexible linker, but the Aar2p-RNaseH complex sequesters the Jab1/MPN domain, thereby preventing binding by Brr2p. Aar2p is phosphorylated in vivo, and a phospho-mimetic S253E mutation in Aar2p leads to disruption of the Aar2p-Prp8p complex in favor of the Brr2p-Prp8p complex. We propose a model in which Aar2p acts as a phosphorylation-controlled U5 snRNP assembly factor that regulates the incorporation of the particle-specific Brr2p. The purpose of this regulation may be to safeguard against nonspecific RNA binding to Prp8p and/or premature activation of Brr2p activity.
关于颗粒特异性蛋白如何在剪接体小核核糖核蛋白（snRNP）上组装，人们知之甚少。Brr2p 是 U5 snRNP 特异性 RNA 解旋酶，对于剪接体催化激活和解体是必需的。在酵母中，Aar2 蛋白是缺乏 Brr2p 的细胞质前体 U5 snRNP 的一部分，在核中被 Brr2p 取代。在这里，我们表明 Aar2p 和 Brr2p 结合到 Prp8p C 末端区域的不同结构域；Aar2p 与核糖核酸酶 H 结构域相互作用，而 Brr2p 与 Jab1/MPN 结构域相互作用。这些结构域通过一个长的、灵活的连接子连接，但 Aar2p-RNaseH 复合物将 Jab1/MPN 结构域隔离，从而阻止 Brr2p 结合。Aar2p 在体内被磷酸化，Aar2p 中的磷酸模拟 S253E 突变导致 Aar2p-Prp8p 复合物的破坏，有利于 Brr2p-Prp8p 复合物的形成。我们提出了一个模型，其中 Aar2p 作为一种磷酸化控制的 U5 snRNP 组装因子，调节颗粒特异性 Brr2p 的掺入。这种调节的目的可能是防止非特异性 RNA 与 Prp8p 结合和/或 Brr2p 活性的过早激活。
