Kv4.2 voltage-gated K+ channel subunits, the primary source of the somatodendritic A-type K+ current in CA1 pyramidal neurons of the hippocampus, play important roles in regulating dendritic excitability and plasticity. To better study the trafficking and subcellular distribution of Kv4.2, we created and characterized a novel Kv4.2 construct encoding a bungarotoxin binding site in the extracellular S3-S4 linker region of the α-subunit. When expressed, this construct can be visualized in living cells after staining with rhodamine-conjugated bungarotoxin. We validated the utility of this construct by visualizing the spontaneous internalization and insertion of Kv4.2 in HEK 293T cells. We further report that Kv4.2 colocalized with several endosome markers in HEK 293T cells. In addition, Kv4.2 internalization is significantly impaired by mitogen-activated protein kinase (MAPK) inhibitors in transfected primary hippocampal neurons. Therefore, this newly developed BBS-Kv4.2 construct provides a novel and powerful tool for studying surface Kv4.2 channel localization and trafficking.

中文翻译：

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一种新型的邦加毒素结合位点标记的结构揭示了MAPK依赖的Kv4.2贩运。

Kv4.2电压门控的K +通道亚基是海马CA1锥体神经元中体树突状A型K +电流的主要来源，在调节树突状兴奋性和可塑性中起重要作用。为了更好地研究Kv4.2的贩运和亚细胞分布，我们创建并表征了一个新颖的Kv4.2构建体，该构建体在α亚基的细胞外S3-S4接头区域中编码一种Bungarotoxin结合位点。当表达时，在用若丹明缀合的邦加毒素染色后，该构建体可以在活细胞中可视化。我们通过可视化HEK 293T细胞中Kv4.2的自发内在化和插入来验证此构建体的实用性。我们进一步报道，Kv4.2与HEK 293T细胞中的几种内体标记共定位。此外，Kv4。在转染的原代海马神经元中，有丝分裂原激活的蛋白激酶（MAPK）抑制剂显着损害了2的内在化。因此，这种新开发的BBS-Kv4.2构造为研究表面Kv4.2通道的定位和运输提供了一个新颖而强大的工具。