The voltage-gated Na⁺ channel β1 subunit, encoded by SCN1B, regulates cell surface expression and gating of α subunits and participates in cell adhesion. β1 is cleaved by α/β and γ-secretases, releasing an extracellular domain and intracellular domain (ICD), respectively. Abnormal SCN1B expression/function is linked to pathologies including epilepsy, cardiac arrhythmia, and cancer. In this study, we sought to determine the effect of secretase cleavage on β1 function in breast cancer cells. Using a series of GFP-tagged β1 constructs, we show that β1-GFP is mainly retained intracellularly, particularly in the endoplasmic reticulum and endolysosomal pathway, and accumulates in the nucleus. Reduction in endosomal β1-GFP levels occurred following γ-secretase inhibition, implicating endosomes and/or the preceding plasma membrane as important sites for secretase processing. Using live-cell imaging, we also report β1ICD-GFP accumulation in the nucleus. Furthermore, β1-GFP and β1ICD-GFP both increased Na⁺ current, whereas β1STOP-GFP, which lacks the ICD, did not, thus highlighting that the β1-ICD is necessary and sufficient to increase Na⁺ current measured at the plasma membrane. Importantly, although the endogenous Na⁺ current expressed in MDA-MB-231 cells is tetrodotoxin (TTX)-resistant (carried by Na_v1.5), the Na⁺ current increased by β1-GFP or β1ICD-GFP was TTX-sensitive. Finally, we found β1-GFP increased mRNA levels of the TTX-sensitive α subunits SCN1A/Na_v1.1 and SCN9A/Na_v1.7. Taken together, this work suggests that the β1-ICD is a critical regulator of α subunit function in cancer cells. Our data further highlight that γ-secretase may play a key role in regulating β1 function in breast cancer.

由SCN1B编码的电压门控 Na ⁺通道 β1 亚基调节细胞表面表达和 α 亚基的门控，并参与细胞粘附。β1 被 α/β 和 γ-分泌酶切割，分别释放胞外结构域和胞内结构域 (ICD)。SCN1B异常表达/功能与包括癫痫、心律失常和癌症在内的病理有关。在这项研究中，我们试图确定分泌酶切割对乳腺癌细胞中 β1 功能的影响。使用一系列 GFP 标记的 β1 构建体，我们表明 β1-GFP 主要保留在细胞内，特别是在内质网和内溶酶体途径中，并在细胞核中积累。γ-分泌酶抑制后发生内体 β1-GFP 水平的降低，暗示内体和/或前面的质膜是分泌酶加工的重要位点。使用活细胞成像，我们还报告了 β1ICD-GFP 在细胞核中的积累。此外，β1-GFP 和 β1ICD-GFP 均增加了 Na ⁺当前，而缺乏 ICD 的 β1STOP-GFP 则没有，因此强调 β1-ICD 是必要且足以增加在质膜上测量的Na ^+电流。重要的是，尽管 MDA-MB-231 细胞中表达的内源性 Na ⁺电流对河豚毒素 (TTX) 具有抗性（由 Na _v 1.5 携带），但 β1-GFP 或 β1ICD-GFP 增加的 Na ⁺电流对 TTX 敏感。最后，我们发现 β1-GFP 增加了 TTX 敏感 α 亚基SCN1A /Na _v 1.1 和SCN9A /Na _{v的 mRNA 水平}1.7. 总之，这项工作表明 β1-ICD 是癌细胞中 α 亚基功能的关键调节剂。我们的数据进一步强调，γ-分泌酶可能在调节乳腺癌中的 β1 功能中发挥关键作用。