Subcellular dynamics and functional activity of the cleaved Na+ channel β1 subunit intracellular domain

Abstract

The voltage-gated Na+ channel {beta}1 subunit, encoded by SCN1B, regulates cell surface expression and gating of subunits, and participates in cell adhesion. {beta}1 is cleaved by /{beta} and {gamma}-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 {beta}1 function in breast cancer cells. Using a series of GFP-tagged {beta}1 constructs, we show that {beta}1-GFP is mainly retained intracellularly, particularly in the endoplasmic reticulum and endolysosomal pathway, and accumulates in the nucleus. Reduction in endosomal {beta}1-GFP levels occurred following {gamma}-secretase inhibition, implicating endosomes, and/or the preceding plasma membrane, as important sites for secretase processing. Using live-cell imaging, we report {beta}1ICD-GFP accumulation in the nucleus. Furthermore, {beta}1-GFP and {beta}1ICD-GFP both increased Na+ current, whereas {beta}1STOP-GFP, which lacks the ICD, did not, thus highlighting that the {beta}1-ICD was 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 TTX-resistant (carried by Nav1.5), the Na+ current increased by {beta}1-GFP or {beta}1ICD-GFP was TTX-sensitive. In addition, {beta}1-GFP increased mRNA levels of the TTX-sensitive subunits SCN1A/Nav1.1 and SCN9A/Nav1.7. Taken together, this work suggests that the {beta}1-ICD is a critical regulator of subunit function in cancer cells. Our data further highlight that {gamma}-secretase may play a key role in regulating {beta}1 function in breast cancer.