Lateral Membrane-Specific MAGUK CASK Down-Regulates NaV1.5 Channel in Cardiac Myocytes
Rationale: Mechanisms underlying membrane protein localization are crucial in the proper function of cardiac myocytes. The main cardiac sodium channel, NaV1.5, carries the sodium current (INa) which provides a rapid depolarizing current during the upstroke of the action potential. While enriched in the intercalated disc (ID), NaV1.5 is present in different membrane domains in myocytes and interacts with several partners.
Objective: To test the hypothesis that the Membrane-Associated GUanylate Kinase (MAGUK) protein Calcium/calmodulin-dependent Serine protein Kinase (CASK) interacts with and regulates NaV1.5 in cardiac myocytes.
Methods and Results: Immunostaining experiments showed that CASK localizes at lateral membranes (LM) of cardiac myocytes, in association with dystrophin. Whole-cell patch clamp showed that CASK-silencing increases in vitro. In vivo CASK knockdown similarly increased INa recorded in freshly isolated myocytes. Pull-down experiments revealed that CASK directly interacts with the C-terminus of NaV1.5. CASK silencing reduces syntrophin expression without affecting NaV1.5 and dystrophin expression levels. Total Internal Reflection Fluorescence microscopy (TIRFm) and biotinylation assays showed that CASK silencing increased the surface expression of NaV1.5 without changing mRNA levels. Quantification of NaV1.5 expression at the LM and ID revealed that the LM pool only was increased upon CASK silencing. The protein transport inhibitor brefeldin-A prevented INa increase in CASK-silenced myocytes. During atrial dilation/remodeling, CASK expression was reduced but its localization unchanged.
Conclusions: This study constitutes the first description of an unconventional MAGUK protein, CASK, which directly interacts with NaV1.5 channel and controls its surface expression at the LM by regulating ion channel trafficking.
- Received June 6, 2016.
- Revision received June 25, 2016.
- Accepted June 30, 2016.