Role of SNAREs and H+-ATPase in the targeting of proton pump-coated vesicles to collecting duct cell apical membrane.
Role of SNAREs and H+-ATPase in the targeting of proton pump-coated vesicles to collecting duct cell apical membrane.
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SNARE 和 H -ATP 酶在质子泵涂层囊泡靶向集合管细胞顶膜中的作用。
DOI：
10.1038/sj.ki.5002500
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发表时间：
2007
影响因子：
19.6
通讯作者：
Alexander,EA
中科院分区：
文献类型：
--
作者：
Schwartz,JH;Li,G;Yang,Q;Suri,V;Ross,JJ;Alexander,EA
H+- atp酶通过囊泡胞吐和胞吞作用向根尖质膜循环，是控制收集管分泌H+的重要机制。我们假设SNAREs（可溶性乙基马来酰亚胺敏感因子附着蛋白）可能参与了H+- atp酶包被囊泡的靶向。利用收集管H+分泌细胞（内髓收集管（IMCD）细胞）的组织培养模型，我们证明了它们表达了snare介导的胞吐所需的蛋白质，并在H+- atp酶胞吐的刺激下形成了snare融合复合物。此外，胞外扩增的根尖H+- atp酶对梭状芽胞杆菌毒素很敏感，这些毒素可以切割SNAREs，从而抑制分泌。因此，SNAREs对于H+- atp酶循环到质膜至关重要。IMCD细胞中的这一过程具有与神经元细胞不同的特征：SNARE复合体包括并需要囊泡货物（H+- atp酶）来靶向。通过对syntaxin 1的嵌合体和截断，我们证明了在syntaxin 1的H3结构域中存在一个特定的盒体，该盒体介导SNAREs的结合，而第二个不同的H3区域则结合H+- atp酶。利用H+- atp酶B1亚基的点突变，我们发现该亚基包含H+- atp酶本身的特定靶向信息。此外，我们发现胞外分泌调节因子Munc-18-2在该系统中发挥多功能作用：它调节SNARE复合物的形成和syntaxin 1对H+-ATPase的亲和力。
Recycling of H+-ATPase to the apical plasma membrane, mediated by vesicular exocytosis and endocytosis, is an important mechanism for controlling H+secretion by the collecting duct. We hypothesized that SNAREs (solubleN-ethylmaleimide-sensitive factor attachment proteins) may be involved in the targeting of H+-ATPase-coated vesicles. Using a tissue culture model of collecting duct H+secretory cells (inner medullary collecting duct (IMCD) cells), we demonstrated that they express the proteins required for SNARE-mediated exocytosis and form SNARE-fusion complexes upon stimulation of H+-ATPase exocytosis. Furthermore, exocytic amplification of apical H+-ATPase is sensitive to clostridial toxins that cleave SNAREs and thereby inhibit secretion. Thus, SNAREs are critical for H+-ATPase cycling to the plasma membrane. The process in IMCD cells has a feature distinct from that of neuronal cells: the SNARE complex includes and requires the vesicular cargo (H+-ATPase) for targeting. Using chimeras and truncations of syntaxin 1, we demonstrated that there is a specific cassette within the syntaxin 1 H3 domain that mediates binding of the SNAREs and a second distinct H3 region that binds H+-ATPase. Utilizing point mutations of the B1 subunit of the H+-ATPase, we document that this subunit contains specific targeting information for the H+-ATPase itself. In addition, we found that Munc-18-2, a regulator of exocytosis, plays a multifunctional role in this system: it regulates SNARE complex formation and the affinity of syntaxin 1 for H+-ATPase.