Research Program in Cell and Molecular Biology, Institute of Biotechnology, University of Helsinki, 00014 Helsinki, Finland.
Mol Biol Cell. 2010 Apr 15;21(8):1362-74. doi: 10.1091/mbc.e09-07-0546. Epub 2010 Feb 24.
Sec1p/Munc18 (SM) family proteins regulate SNARE complex function in membrane fusion through their interactions with syntaxins. In addition to syntaxins, only a few SM protein interacting proteins are known and typically, their binding modes with SM proteins are poorly characterized. We previously identified Mso1p as a Sec1p-binding protein and showed that it is involved in membrane fusion regulation. Here we demonstrate that Mso1p and Sec1p interact at sites of exocytosis and that the Mso1p-Sec1p interaction site depends on a functional Rab GTPase Sec4p and its GEF Sec2p. Random and targeted mutagenesis of Sec1p, followed by analysis of protein interactions, indicates that Mso1p interacts with Sec1p domain 1 and that this interaction is important for membrane fusion. In many SM family proteins, domain 1 binds to a N-terminal peptide of a syntaxin family protein. The Sec1p-interacting syntaxins Sso1p and Sso2p lack the N-terminal peptide. We show that the putative N-peptide binding area in Sec1p domain 1 is important for Mso1p binding, and that Mso1p can interact with Sso1p and Sso2p. Our results suggest that Mso1p mimics N-peptide binding to facilitate membrane fusion.
Sec1p/Munc18（SM）家族蛋白通过与突触融合蛋白（syntaxin）的相互作用来调节 SNARE 复合物的功能。除了突触融合蛋白之外，目前仅知道少数几个与 SM 蛋白相互作用的蛋白质，而且通常，它们与 SM 蛋白的结合模式也没有得到很好的描述。我们之前鉴定出 Mso1p 是一种与 Sec1p 结合的蛋白，并表明它参与了膜融合的调节。在这里，我们证明 Mso1p 和 Sec1p 在胞吐作用的部位相互作用，并且 Mso1p-Sec1p 相互作用位点依赖于功能性 Rab GTPase Sec4p 和其 GEF Sec2p。对 Sec1p 进行随机和靶向诱变，然后分析蛋白质相互作用，表明 Mso1p 与 Sec1p 结构域 1 相互作用，并且这种相互作用对膜融合很重要。在许多 SM 家族蛋白中，结构域 1 与突触融合蛋白家族蛋白的 N 端肽结合。与 Sec1p 相互作用的突触融合蛋白 Sso1p 和 Sso2p 缺乏 N 端肽。我们表明，Sec1p 结构域 1 中的假定 N-肽结合区域对 Mso1p 结合很重要，并且 Mso1p 可以与 Sso1p 和 Sso2p 相互作用。我们的结果表明，Mso1p 模拟 N-肽结合以促进膜融合。
