Max Perutz Labs, Vienna Biocenter, Medical University of Vienna, Vienna, Austria.
Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, United States.
Elife. 2022 Aug 18;11:e82041. doi: 10.7554/eLife.82041.
Exocytosis is an active vesicle trafficking process by which eukaryotes secrete materials to the extracellular environment and insert membrane proteins into the plasma membrane. The final step of exocytosis in yeast involves the assembly of two t-SNAREs, Sso1/2 and Sec9, with the v-SNARE, Snc1/2, on secretory vesicles. The rate-limiting step in this process is the formation of a binary complex of the two t-SNAREs. Despite a previous report of acceleration of binary complex assembly by Sec3, it remains unknown how Sso2 is efficiently recruited to the vesicle-docking site marked by Sec3. Here, we report a crystal structure of the pleckstrin homology (PH) domain of Sec3 in complex with a nearly full-length version of Sso2 lacking only its C-terminal transmembrane helix. The structure shows a previously uncharacterized binding site for Sec3 at the N-terminus of Sso2, consisting of two highly conserved triple residue motifs (NPY: Asn-Pro-Tyr). We further reveal that the two NPY motifs bind Sec3 synergistically, which together with the previously reported binding interface constitute dual-site interactions between Sso2 and Sec3 to drive the fusion of secretory vesicles at target sites on the plasma membrane.
胞吐作用是真核生物将物质分泌到细胞外环境并将膜蛋白插入质膜的一种主动囊泡运输过程。在酵母中，胞吐作用的最后一步涉及到两个 t-SNARE，Sso1/2 和 Sec9，与囊泡上的 v-SNARE，Snc1/2 的组装。这个过程的限速步骤是两个 t-SNARE 形成二元复合物。尽管之前有报道称 Sec3 可以加速二元复合物的组装，但 Sso2 如何有效地被招募到由 Sec3 标记的囊泡停泊位点仍然未知。在这里，我们报告了 Sec3 的pleckstrin 同源（PH）结构域与缺少其 C 末端跨膜螺旋的全长 Sso2 的复合物的晶体结构。该结构显示了 Sec3 在 Sso2 的 N 末端的一个以前未被描述的结合位点，由两个高度保守的三氨基酸基序（NPY：天冬酰胺-脯氨酸-酪氨酸）组成。我们进一步揭示了两个 NPY 基序协同结合 Sec3，这与之前报道的结合界面一起构成了 Sso2 和 Sec3 之间的双位点相互作用，以驱动分泌囊泡在质膜上目标位点的融合。
