VIB Center for the Biology of Disease, 3000 Leuven, Belgium.
Neuron. 2013 Mar 20;77(6):1097-108. doi: 10.1016/j.neuron.2013.01.025.
PI(3,4,5)P3 is a low-abundance lipid thought to play a role in the regulation of synaptic activity; however, the mechanism remains obscure. We have constructed novel split Venus-based probes and used superresolution imaging to localize PI(3,4,5)P3 at Drosophila larval neuromuscular synapses. We find the lipid in membrane domains at neurotransmitter release sites, where it concentrates with Syntaxin1A, a protein essential for vesicle fusion. Reducing PI(3,4,5)P3 availability disperses Syntaxin1A clusters and increasing PI(3,4,5)P3 levels rescues this defect. In artificial giant unilamellar vesicles, PI(3,4,5)P3 also induces Syntaxin1A domain formation and this clustering, in vitro and in vivo, is dependent on positively charged residues in the Syntaxin1A-juxtamembrane domain. Functionally, reduced PI(3,4,5)P3 causes temperature-sensitive paralysis and reduced neurotransmitter release, a phenotype also seen in animals expressing a Syntaxin1A with a mutated juxtamembrane domain. Thus, our data indicate that PI(3,4,5)P3, based on electrostatic interactions, clusters Syntaxin1A at release sites to regulate neurotransmitter release.
PI(3,4,5)P3 是一种低丰度脂质，被认为在调节突触活动中发挥作用；然而，其机制尚不清楚。我们构建了新型的 Venus 分割探针，并利用超分辨率成像技术在果蝇幼虫神经肌肉突触上定位了 PI(3,4,5)P3。我们发现脂质存在于神经递质释放位点的膜域中，在这些位点与 Syntaxin1A 聚集，Syntaxin1A 是囊泡融合所必需的一种蛋白质。减少 PI(3,4,5)P3 的可用性会分散 Syntaxin1A 簇，而增加 PI(3,4,5)P3 水平则可以挽救这种缺陷。在人工巨单层囊泡中，PI(3,4,5)P3 也诱导了 Syntaxin1A 结构域的形成，这种体外和体内的聚类依赖于 Syntaxin1A 跨膜结构域中的正电荷残基。在功能上，减少 PI(3,4,5)P3 会导致热敏性麻痹和神经递质释放减少，这种表型也见于表达具有突变跨膜结构域的 Syntaxin1A 的动物中。因此，我们的数据表明，基于静电相互作用，PI(3,4,5)P3 在释放位点聚集 Syntaxin1A 以调节神经递质释放。
