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Reconstitution of Rab- and SNARE-dependent membrane fusion by synthetic endosomes

机译：合成内体重建Rab和SNARE依赖性膜融合

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摘要

Rab GTPases and SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) are evolutionarily conserved essential components of the eukaryotic intracellular transport system. Although pairing of cognate SNAREs is sufficient to fuse membranes in vitro, a complete reconstitution of the Rab-SNARE machinery has never been achieved. Here we report the reconstitution of the early endosomal canine Rab5 GTPase, its key regulators and effectors together with SNAREs into proteoliposomes using a set of 17 recombinant human proteins. These vesicles behave like minimal 'synthetic' endosomes, fusing with purified early endosomes or with each other in vitro. Membrane fusion measured by content-mixing and morphological assays requires the cooperativity between Rab5 effectors and cognate SNAREs which, together, form a more efficient 'core machinery' than SNAREs alone. In reconstituting a fusion mechanism dependent on both a Rab GTPase and SNAREs, our work shows that the two machineries act coordinately to increase the specificity and efficiency of thernmembrane tethering and fusion process.
机译：Rab GTPases和SNARE（可溶性N-乙基马来酰亚胺敏感因子附着蛋白受体）是真核细胞内转运系统的进化保守组成部分。尽管配对的SNARE配对足以在体外融合膜，但Rab-SNARE机械的完全重组从未实现。在这里，我们报告使用一组17种重组人蛋白将早期的内体犬Rab5 GTPase，其关键调节剂和效应子与SNARE一起重建为脂质体。这些囊泡的行为就像最小的“合成”内体，与纯化的早期内体融合或在体外彼此融合。通过内容混合和形态学测定法测量的膜融合需要Rab5效应子与同源SNARE之间的协同作用，它们共同构成比单独的SNARE更有效的“核心机制”。在重构依赖于Rab GTPase和SNARE的融合机制时，我们的工作表明，这两种机制协同作用以提高膜束缚和融合过程的特异性和效率。

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  • 来源
    《Nature》 |2009年第25期|1091-1097|共7页
  • 作者

    Takeshi Ohya; Marta Miaczynska; UEnal Coskun; Barbara Lommer; Anja Runge; David Drechsel; Yannis Kalaidzidis; Marino Zerial;

  • 作者单位

    Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01309, Dresden, Germany.;

    Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01309, Dresden, Germany.;

    Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01309, Dresden, Germany.;

    Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01309, Dresden, Germany.;

    Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01309, Dresden, Germany.;

    Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01309, Dresden, Germany.;

    Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01309, Dresden, Germany A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow 119899, Russia. fPresent address: International Institute of Molecular and Cell Biology (IIMCB), Ks. Trojdena 4, 02-109 Warsaw, Poland;

    Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstrasse 108, 01309, Dresden, Germany.;

  • 收录信息 美国《科学引文索引》(SCI);美国《工程索引》(EI);美国《生物学医学文摘》(MEDLINE);美国《化学文摘》(CA);
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