Interaction between the RNA binding domains of Ser-Arg splicing factor 1 and U1-70K snRNP protein determines early spliceosome assembly
Interaction between the RNA binding domains of Ser-Arg splicing factor 1 and U1-70K snRNP protein determines early spliceosome assembly
复制标题
DOI：
10.1073/pnas.1017700108
复制
发表时间：
2011-05-17
影响因子：
11.1
通讯作者：
Ghosh, Gourisankar
中科院分区：
文献类型：
--
作者：
Cho, Suhyung;Hoang, Amy;Ghosh, Gourisankar
在哺乳动物细胞中，剪接体的早期组装是由U1小核核糖核蛋白（U1 snRNP）结合到5'剪接位点（5' SS）开始的，并由富含Ser/Arg（SR）的蛋白质辅助。在这个过程中，SR蛋白的RS结构域被认为与U1- 70 K的RS基序直接相互作用，而U1- 70 K的RS基序受到RS结构域磷酸化的调节。在这里，我们报告的早期剪接体组装事件是由丝氨酸/丝氨酸丰富的剪接因子1（SRSF 1）的RNA识别域（RRM）介导的，它桥的RRM的U1- 70 K的前mRNA通过使用表面相反的RNA结合位点。SRSF 1的RRM中的特异性突变破坏RRM-RRM相互作用也抑制剪接体E复合物的形成和剪接。我们进一步证明，低磷酸化的RS结构域的SRSF 1与自己的RRM相互作用，从而与U1- 70 K的结合竞争，而超磷酸化的RS结构域允许形成一个三元复合物，包含ESE，SR蛋白，和U1 snRNP。因此，SRSF 1中RS结构域的磷酸化似乎诱导了从分子内相互作用到分子间相互作用的关键分子开关，这表明在前体mRNA剪接期间记录的磷酸化/去磷酸化循环的要求是合理的机制。
It has been widely accepted that the early spliceosome assembly begins with U1 small nuclear ribonucleoprotein (U1 snRNP) binding to the 5' splice site (5' SS), which is assisted by the Ser/Arg (SR)-rich proteins in mammalian cells. In this process, the RS domain of SR proteins is thought to directly interact with the RS motif of U1-70K, which is subject to regulation by RS domain phosphorylation. Here we report that the early spliceosome assembly event is mediated by the RNA recognition domains (RRM) of serine/arginine-rich splicing factor 1 (SRSF1), which bridges the RRM of U1-70K to pre-mRNA by using the surface opposite to the RNA binding site. Specific mutation in the RRM of SRSF1 that disrupted the RRM-RRM interaction also inhibits the formation of spliceosomal E complex and splicing. We further demonstrate that the hypo-phosphorylated RS domain of SRSF1 interacts with its own RRM, thus competing with U1-70K binding, whereas the hyper-phosphorylated RS domain permits the formation of a ternary complex containing ESE, an SR protein, and U1 snRNP. Therefore, phosphorylation of the RS domain in SRSF1 appears to induce a key molecular switch from intra-to intermolecular interactions, suggesting a plausible mechanism for the documented requirement for the phosphorylation/dephosphorylation cycle during pre-mRNA splicing.