Genetic and functional studies have confirmed an important role for the voltage-gated sodium channel Nav1.9 in human pain disorders. However, low functional expression of Nav1.9 in heterologous systems (e.g. in human embryonic kidney 293 (HEK293) cells) has hampered studies of its biophysical and pharmacological properties and the development of high-throughput assays for drug development targeting this channel. The mechanistic basis for the low level of Nav1.9 currents in heterologous expression systems is not understood. Here, we implemented a multidisciplinary approach to investigate the mechanisms that govern functional Nav1.9 expression. Recombinant expression of a series of Nav1.9-Nav1.7 C-terminal chimeras in HEK293 cells identified a 49-amino-acid-long motif in the C terminus of the two channels that regulates expression levels of these chimeras. We confirmed the critical role of this motif in the context of a full-length channel chimera, Nav1.9-Ct49aaNav1.7, which displayed significantly increased current density in HEK293 cells while largely retaining the characteristic Nav1.9-gating properties. High-resolution live microscopy indicated that the newly identified C-terminal motif dramatically increases the number of channels on the plasma membrane of HEK293 cells. Molecular modeling results suggested that this motif is exposed on the cytoplasmic face of the folded C terminus, where it might interact with other channel partners. These findings reveal that a 49-residue-long motif in Nav1.9 regulates channel trafficking to the plasma membrane.

中文翻译：

---

Nav1.9的C末端有49个残基的序列基序调节通道向质膜的运输。

遗传和功能研究已证实，电压门控钠通道Nav1.9在人类疼痛疾病中具有重要作用。但是，Nav1.9在异源系统（例如在人类胚胎肾293（HEK293）细胞中）的低功能表达阻碍了其生物物理和药理学特性以及针对该通道的药物开发高通量分析方法的研究。不了解异源表达系统中Nav1.9电流低水平的机理基础。在这里，我们实现了一种多学科的方法来研究控制功能性Nav1.9表达的机制。一系列Nav1.9-Nav1的重组表达。HEK293细胞中的7个C末端嵌合体在两个通道的C末端鉴定了一个49个氨基酸长的基序，该基序调节了这些嵌合体的表达水平。我们在全长通道嵌合体Nav1.9-Ct49aaNav1.7的背景下证实了该基序的关键作用，该嵌合体在HEK293细胞中显示出显着增加的电流密度，同时很大程度上保留了Nav1.9门控特性。高分辨率现场显微镜检查表明，新近鉴定出的C末端基序显着增加了HEK293细胞质膜上的通道数量。分子建模结果表明，该基序暴露在折叠的C末端的胞质面上，在此处可能与其他通道伴侣相互作用。这些发现揭示了Nav1中有49个残基长的基序。