Kv2.1 cell surface clusters are insertion platforms for ion channel delivery to the plasma membrane.
Kv2.1 cell surface clusters are insertion platforms for ion channel delivery to the plasma membrane.
复制标题
DOI：
10.1091/mbc.e12-01-0047
复制
发表时间：
2012-08
影响因子：
3.3
通讯作者：
Tamkun MM
中科院分区：
文献类型：
--
作者：
Deutsch E;Weigel AV;Akin EJ;Fox P;Hansen G;Haberkorn CJ;Loftus R;Krapf D;Tamkun MM
转基因HEK细胞和海马神经元中的Kv2.1表面簇被证明是参与钾通道进出细胞表面的转运平台。这项工作首次定义了稳定的细胞表面位置，用于离子通道在细胞表面的传递和检索。电压门控K+(Kv)通道在许多细胞类型中调节膜电位。尽管必须很好地控制通道表面的密度和位置，但对Kv通道在细胞表面的传递和恢复知之甚少。Kv2.1通道定位于神经元和转染人胚胎肾(HEK)细胞中微米级的簇，在那里它是不传导的。由于Kv2.1被认为参与了可溶性N-乙基马来酰亚胺敏感因子结合蛋白受体介导的膜融合，因此我们验证了这些表面簇是参与膜蛋白运输的专门平台的假设。对单个Kv2.1通道进行光漂白和量子点成像研究后，基于全内反射的荧光恢复显示，在转染的HEK细胞和海马神经元中，含有Kv2.1的囊泡在Kv2.1表面簇传递货物。在两种细胞中，超过85%的细胞质和循环Kv2.1通道被输送到簇周的细胞表面。与Kv2.1不同，至少85%的回收Kv1.4也是在这里交付的，因为Kv2.1的表面分布均匀。肌动蛋白解聚导致Kv2.1在无簇状表面膜上胞吐。这些结果表明，Kv2.1的一个非传导功能是形成参与膜蛋白运输的微区。这项研究是第一次确定参与离子通道运输的稳定细胞表面平台。
Kv2.1 surface clusters in transfected HEK cells and hippocampal neurons are shown to be trafficking platforms involved in potassium channel movement to and from the cell surface. This work is the first to define stable cell surface sites for ion channel delivery and retrieval at the cell surface. Voltage-gated K+ (Kv) channels regulate membrane potential in many cell types. Although the channel surface density and location must be well controlled, little is known about Kv channel delivery and retrieval on the cell surface. The Kv2.1 channel localizes to micron-sized clusters in neurons and transfected human embryonic kidney (HEK) cells, where it is nonconducting. Because Kv2.1 is postulated to be involved in soluble N-ethylmaleimide–sensitive factor attachment protein receptor–mediated membrane fusion, we examined the hypothesis that these surface clusters are specialized platforms involved in membrane protein trafficking. Total internal reflection–based fluorescence recovery after photobleaching studies and quantum dot imaging of single Kv2.1 channels revealed that Kv2.1-containing vesicles deliver cargo at the Kv2.1 surface clusters in both transfected HEK cells and hippocampal neurons. More than 85% of cytoplasmic and recycling Kv2.1 channels was delivered to the cell surface at the cluster perimeter in both cell types. At least 85% of recycling Kv1.4, which, unlike Kv2.1, has a homogeneous surface distribution, is also delivered here. Actin depolymerization resulted in Kv2.1 exocytosis at cluster-free surface membrane. These results indicate that one nonconducting function of Kv2.1 is to form microdomains involved in membrane protein trafficking. This study is the first to identify stable cell surface platforms involved in ion channel trafficking.