Multivalent Rab interactions determine tether-mediated membrane fusion.
Multivalent Rab interactions determine tether-mediated membrane fusion.
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DOI：
10.1091/mbc.e16-11-0764
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发表时间：
2017-01-15
影响因子：
3.3
通讯作者：
Ungermann C
中科院分区：
文献类型：
--
作者：
Lürick A;Gao J;Kuhlee A;Yavavli E;Langemeyer L;Perz A;Raunser S;Ungermann C
HOPS 束缚复合物结合 Rab7 样 Ypt7 和 SNARE。几个 HOPS 突变体用于表明，Rab 结合位点（而非 Vps41 中的 ALPS 基序）对于束缚和融合膜是必需的。内膜处的膜融合需要 Rab GTPases 和系链之间的串扰来驱动 SNARE 介导的脂质双层混合。一些系链具有多个 Rab 结合位点，其功能很大程度上未经测试。在这里，我们将溶酶体 HOPS 复合物剖析为一种束缚复合物，仅具有两个 Rab7 样 Ypt7 蛋白的结合位点，以确定它们与融合的相关性。使用与 HOPS 突变体相结合的束缚和融合测定，我们表明 HOPS 依赖性融合需要两个 Rab 结合位点，其中 Vps39 是比 Vps41 更强的 Ypt7 相互作用子。 HOPS Vps41 内的内在两亲性脂质包装传感器 (ALPS) 基序是液泡激酶 Yck3 的靶标，对于束缚和融合来说是可有可无的，但如果磷酸化，可能会影响束缚。结合起来，我们的数据表明，多价束缚复合物使用其两个 Rab 结合来确定 SNARE 组装的位置，从而确定内膜上的融合。
The HOPS tethering complex binds both the Rab7-like Ypt7 and SNAREs. Several HOPS mutants are used to show that both Rab-binding sites, but not the ALPS motif in Vps41, are necessary to tether and fuse membranes. Membrane fusion at endomembranes requires cross-talk between Rab GTPases and tethers to drive SNARE-mediated lipid bilayer mixing. Several tethers have multiple Rab-binding sites with largely untested function. Here we dissected the lysosomal HOPS complex as a tethering complex with just two binding sites for the Rab7-like Ypt7 protein to determine their relevance for fusion. Using tethering and fusion assays combined with HOPS mutants, we show that HOPS-dependent fusion requires both Rab-binding sites, with Vps39 being the stronger Ypt7 interactor than Vps41. The intrinsic amphipathic lipid packaging sensor (ALPS) motif within HOPS Vps41, a target of the vacuolar kinase Yck3, is dispensable for tethering and fusion but can affect tethering if phosphorylated. In combination, our data demonstrate that a multivalent tethering complex uses its two Rab bindings to determine the place of SNARE assembly and thus fusion at endomembranes.