D'Agostino Massimo, Risselada Herre Jelger, Mayer Andreas
Département de Biochimie, Université de Lausanne, Epalinges, Switzerland.
Department of Theoretical Physics, Georg-August University, Goettingen, Germany.
EMBO Rep. 2016 Nov;17(11):1590-1608. doi: 10.15252/embr.201642209. Epub 2016 Sep 19.
SNAREs fuse membranes in several steps. Trans-SNARE complexes juxtapose membranes, induce hemifused stalk structures, and open the fusion pore. A recent penetration model of fusion proposed that SNAREs force the hydrophilic C-termini of their transmembrane domains through the hydrophobic core of the membrane(s). In contrast, the indentation model suggests that the C-termini open the pore by locally compressing and deforming the stalk. Here we test these models in the context of yeast vacuole fusion. Addition of small hydrophilic tags renders bilayer penetration by the C-termini energetically unlikely. It preserves fusion activity, however, arguing against the penetration model. Addition of large protein tags to the C-termini permits SNARE activation, trans-SNARE pairing, and hemifusion but abolishes pore opening. Fusion proceeds if the tags are detached from the membrane by a hydrophilic spacer or if only one side of the trans-SNARE complex carries a protein tag. Thus, both sides of a trans-SNARE complex can drive pore opening. Our results are consistent with an indentation model in which multiple SNARE C-termini cooperate in opening the fusion pore by locally deforming the inner leaflets.
可溶性N-乙基马来酰胺敏感因子附着蛋白受体（SNAREs）通过几个步骤使膜融合。跨SNARE复合物使膜并列，诱导半融合茎状结构，并打开融合孔。最近提出的一种融合渗透模型认为，SNAREs迫使它们跨膜结构域的亲水性C末端穿过膜的疏水核心。相比之下，压痕模型表明，C末端通过局部压缩和使茎状结构变形来打开孔。在这里，我们在酵母液泡融合的背景下测试这些模型。添加小的亲水性标签使C末端在能量上不太可能穿透双层膜。然而，它保留了融合活性，这与渗透模型相矛盾。在C末端添加大的蛋白质标签允许SNARE激活、跨SNARE配对和半融合，但会消除孔的打开。如果标签通过亲水性间隔物与膜分离，或者如果跨SNARE复合物只有一侧带有蛋白质标签，则融合会继续进行。因此，跨SNARE复合物的两侧都可以驱动孔的打开。我们的结果与压痕模型一致，在该模型中，多个SNARE C末端通过局部使内膜小叶变形来协同打开融合孔。
