Institute of Biochemistry, Department of Biology, ETH Zürich CH-8093 Zürich, Switzerland.
Department of Basic Medical Sciences, University of Arizona, College of Medicine-Phoenix, Phoenix, AZ 85004.
Proc Natl Acad Sci U S A. 2022 Feb 8;119(6). doi: 10.1073/pnas.2114092119.
In mammals, the structural basis for the interaction between U1 and U2 small nuclear ribonucleoproteins (snRNPs) during the early steps of splicing is still elusive. The binding of the ubiquitin-like (UBL) domain of SF3A1 to the stem-loop 4 of U1 snRNP (U1-SL4) contributes to this interaction. Here, we determined the 3D structure of the complex between the UBL of SF3A1 and U1-SL4 RNA. Our crystallography, NMR spectroscopy, and cross-linking mass spectrometry data show that SF3A1-UBL recognizes, sequence specifically, the GCG/CGC RNA stem and the apical UUCG tetraloop of U1-SL4. In vitro and in vivo mutational analyses support the observed intermolecular contacts and demonstrate that the carboxyl-terminal arginine-glycine-glycine-arginine (RGGR) motif of SF3A1-UBL binds sequence specifically by inserting into the RNA major groove. Thus, the characterization of the SF3A1-UBL/U1-SL4 complex expands the repertoire of RNA binding domains and reveals the capacity of RGG/RG motifs to bind RNA in a sequence-specific manner.
在哺乳动物中，U1 和 U2 小核核糖核蛋白（snRNP）在剪接早期步骤中相互作用的结构基础仍然难以捉摸。SF3A1 的泛素样（UBL）结构域与 U1 snRNP 的茎环 4（U1-SL4）的结合有助于这种相互作用。在这里，我们确定了 SF3A1 的 UBL 与 U1-SL4 RNA 之间复合物的 3D 结构。我们的晶体学、NMR 光谱学和交联质谱数据表明，SF3A1-UBL 特异性识别 GCG/CGC RNA 茎和 U1-SL4 的顶端 UUCG 四联体。体外和体内突变分析支持观察到的分子间接触，并证明 SF3A1-UBL 的羧基末端精氨酸-甘氨酸-甘氨酸-精氨酸（RGGR）基序通过插入 RNA 大沟特异性结合序列。因此，SF3A1-UBL/U1-SL4 复合物的表征扩展了 RNA 结合结构域的范围，并揭示了 RGG/RG 基序以序列特异性方式结合 RNA 的能力。
