Otto-Warburg Laboratory, Max-Planck Institute for Molecular Genetics, 14195 Berlin, Germany.
Mol Cell. 2012 Feb 24;45(4):567-80. doi: 10.1016/j.molcel.2011.12.034.
More than 200 proteins copurify with spliceosomes, the compositionally dynamic RNPs catalyzing pre-mRNA splicing. To better understand protein - protein interactions governing splicing, we systematically investigated interactions between human spliceosomal proteins. A comprehensive Y2H interaction matrix screen generated a protein interaction map comprising 632 interactions between 196 proteins. Among these, 242 interactions were found between spliceosomal core proteins and largely validated by coimmunoprecipitation. To reveal dynamic changes in protein interactions, we integrated spliceosomal complex purification information with our interaction data and performed link clustering. These data, together with interaction competition experiments, suggest that during step 1 of splicing, hPRP8 interactions with SF3b proteins are replaced by hSLU7, positioning this second step factor close to the active site, and that the DEAH-box helicases hPRP2 and hPRP16 cooperate through ordered interactions with GPKOW. Our data provide extensive information about the spliceosomal protein interaction network and its dynamics.
超过 200 种蛋白质与剪接体共纯化，剪接体是催化前体 mRNA 剪接的组成动态 RNA 蛋白复合物。为了更好地理解剪接过程中的蛋白质-蛋白质相互作用，我们系统地研究了人类剪接体蛋白之间的相互作用。全面的 Y2H 相互作用矩阵筛选产生了一个包含 196 种蛋白质之间 632 种相互作用的蛋白质相互作用图谱。其中，242 种相互作用存在于剪接体核心蛋白之间，并通过共免疫沉淀得到了很大程度的验证。为了揭示蛋白质相互作用的动态变化，我们将剪接体复合物纯化信息与我们的相互作用数据进行了整合，并进行了链接聚类。这些数据以及相互作用竞争实验表明，在剪接的第一步中，hPRP8 与 SF3b 蛋白的相互作用被 hSLU7 取代，将第二步因子定位到活性位点附近，并且 DEAH 盒解旋酶 hPRP2 和 hPRP16 通过与 GPKOW 的有序相互作用进行合作。我们的数据提供了关于剪接体蛋白相互作用网络及其动态的广泛信息。
