Kiessling Volker, Liang Binyong, Tamm Lukas K
Center for Membrane Biology and Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, VA, USA.
Methods Cell Biol. 2015;128:339-63. doi: 10.1016/bs.mcb.2015.02.005. Epub 2015 Apr 8.
Successful reconstitutions of SNARE-mediated intracellular membrane fusion have been achieved in bulk fusion assays since 1998 and in single liposome fusion assays since 2004. Especially in neuronal presynaptic SNARE-mediated exocytosis, fusion is controlled by numerous accessory proteins, of which some functions have also been reconstituted in vitro. The development of and results obtained with two fundamentally different single liposome fusion assays, namely liposome-to-supported membrane and liposome-to-liposome, are reviewed. Both assays distinguish between liposome docking and fusion steps of the overall fusion reaction and both assays are capable of resolving hemi-and full-fusion intermediates and end states. They have opened new windows for elucidating the mechanisms of these fundamentally important cellular reactions with unprecedented time and molecular resolution. Although many of the molecular actors in this process have been discovered, we have only scratched the surface of looking at their fascinating plays, interactions, and choreographies that lead to vesicle traffic as well as neurotransmitter and hormone release in the cell.
自1998年以来，在大量融合试验中成功实现了SNARE介导的细胞内膜融合，自2004年以来在单脂质体融合试验中也实现了这一目标。特别是在神经元突触前SNARE介导的胞吐作用中，融合受多种辅助蛋白控制，其中一些功能也已在体外重建。本文综述了两种根本不同的单脂质体融合试验，即脂质体与支持膜融合试验和脂质体与脂质体融合试验的发展情况及所取得的结果。这两种试验都区分了整体融合反应中的脂质体对接和融合步骤，并且都能够分辨半融合和完全融合中间体以及最终状态。它们以前所未有的时间和分子分辨率为阐明这些至关重要的细胞反应机制打开了新的窗口。尽管在这个过程中已经发现了许多分子参与者，但我们对于它们在导致细胞内囊泡运输以及神经递质和激素释放的过程中所展现出的迷人作用、相互作用和编排，才刚刚触及皮毛。
