Department of Chemistry & Biochemistry, University of California, San Diego 9500 Gilman Drive, La Jolla, CA 92093, USA.
Nucleic Acids Res. 2011 Nov;39(21):9413-21. doi: 10.1093/nar/gkr663. Epub 2011 Aug 18.
SR proteins promote spliceosome formation by recognizing exonic splicing enhancers (ESEs) during pre-mRNA splicing. Each SR protein binds diverse ESEs using strategies that are yet to be elucidated. Here, we show that the RNA-binding domain (RBD) of SRSF1 optimally binds to decameric purine rich ESE sequences although locations of purines are not stringently specified. The presence of uracils either within or outside of the recognition site is detrimental for binding with SRSF1. The entire RBD, comprised of two RRMs and a glycine-rich linker, is essential for ESE binding. Mutation within each segment reduced or nearly abolished binding, suggesting that these segments mediate cooperative binding. The linker plays a decisive role in organizing ESE binding. The flanking basic regions of the linker appear to communicate with each other in bringing the two RRMs close together to form the complex with RNA. Our study thus suggests semi-conservative adaptable interaction between ESE and SRSF1, and such binding mode is not only essential for the recognition of plethora of physiological ESE sequences but may also be essential for the interaction with various factors during the spliceosome assembly.
SR 蛋白通过在 pre-mRNA 剪接过程中识别外显子剪接增强子 (ESE) 来促进剪接体的形成。每个 SR 蛋白都使用尚未阐明的策略结合不同的 ESE。在这里，我们表明，SRSF1 的 RNA 结合结构域 (RBD) 最优化地结合到包含十个嘧啶的富含嘌呤的 ESE 序列，尽管嘧啶的位置没有严格指定。嘧啶无论是在识别位点内还是外，都会对与 SRSF1 的结合产生不利影响。由两个 RRM 和一个甘氨酸丰富的连接子组成的整个 RBD 对于 ESE 结合是必需的。每个片段内的突变减少或几乎消除了结合，这表明这些片段介导协同结合。连接子在组织 ESE 结合方面起着决定性的作用。连接子的侧翼碱性区域似乎在彼此之间进行通信，使两个 RRM 紧密结合，从而与 RNA 形成复合物。因此，我们的研究表明，ESE 和 SRSF1 之间存在半保守适应性相互作用，这种结合模式不仅对于识别大量生理 ESE 序列是必需的，而且对于在剪接体组装过程中与各种因子相互作用也是必需的。
