The U11/U12 snRNP 65K protein acts as a molecular bridge, binding the U12 snRNA and U11-59K protein
The U11/U12 snRNP 65K protein acts as a molecular bridge, binding the U12 snRNA and U11-59K protein
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DOI：
10.1038/sj.emboj.7600765
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发表时间：
2005-09-07
期刊：
影响因子：
11.4
通讯作者：
Will, CL
中科院分区：
文献类型：
--
作者：
Benecke, H;Lührmann, R;Will, CL
U11和U12与前mRNA的50剪接位点和分支位点协同作用，形成稳定的双snRNP复合物，从而桥接U12依赖性前剪接体内内含子的5'和3'末端。为了鉴定有助于双snRNP形成和内含子桥接的蛋白质，我们研究了U11/ U12 snRNP组分之间的蛋白质-蛋白质和蛋白质- RNA相互作用。我们证明了U11/ U12- 65 K蛋白具有双重结合活性，通过其C-末端RRM直接与U12 snRNA相互作用，通过其N-末端的一半与U11相关的59 K蛋白相互作用。我们提供的证据表明，与以前发表的U12 snRNA二级结构模型相比，U12的30半形成了一个高度保守的7个核苷酸环的延长茎环，后者作为65 K结合位点。在体外剪接反应中添加包含65 K结合位点的寡核苷酸抑制U12依赖性，但不抑制U2依赖性的前mRNA剪接。综上所述，这些数据表明U11/ U12- 65 K和U11- 59 K有助于小前剪接体中的双snRNP形成和内含子桥接。
U11 and U12 interact cooperatively with the 50 splice site and branch site of pre-mRNA as a stable preformed di-snRNP complex, thereby bridging the 5' and 3' ends of the intron within the U12-dependent prespliceosome. To identify proteins contributing to di-snRNP formation and intron bridging, we investigated protein - protein and protein - RNA interactions between components of the U11/ U12 snRNP. We demonstrate that the U11/ U12-65K protein possesses dual binding activity, interacting directly with U12 snRNA via its C-terminal RRM and the U11-associated 59K protein via its N-terminal half. We provide evidence that, in contrast to the previously published U12 snRNA secondary structure model, the 30 half of U12 forms an extended stem-loop with a highly conserved seven-nucleotide loop and that the latter serves as the 65K binding site. Addition of an oligonucleotide comprising the 65K binding site to an in vitro splicing reaction inhibited U12-dependent, but not U2-dependent, pre-mRNA splicing. Taken together, these data suggest that U11/ U12-65K and U11-59K contribute to di-snRNP formation and intron bridging in the minor prespliceosome.