Divergent regulation of KCNQ1/E1 by targeted recruitment of protein kinase A to distinct sites on the channel complex
- Columbia University, United States
Abstract
The slow delayed rectifier potassium current, IKs, conducted through pore-forming Q1 and auxiliary E1 ion channel complexes is important for human cardiac action potential repolarization. During exercise or fright, IKs is up-regulated by protein kinase A (PKA)-mediated Q1 phosphorylation to maintain heart rhythm and optimum cardiac performance. Sympathetic upregulation of IKs requires recruitment of PKA holoenzyme (two regulatory- RI or RII- and two catalytic Cα subunits) to Q1 C-terminus by an A kinase anchoring protein (AKAP9). Mutations in Q1 or AKAP9 that abolish their functional interaction result in long QT syndrome type 1 and 11, respectively, which increases the risk of sudden cardiac death during exercise. Here, we investigated the utility of a targeted protein phosphorylation (TPP) approach to reconstitute PKA regulation of IKs in the absence of AKAP9. Targeted recruitment of endogenous Cα to E1-YFP using a GFP/YFP nanobody (nano) fused to RIIα enabled acute cAMP-mediated enhancement of IKs, reconstituting physiological regulation of the channel complex. By contrast, nano-mediated tethering of RIIα or Cα to Q1-YFP constitutively inhibited IKs by retaining the channel intracellularly in the endoplasmic reticulum and Golgi. Proteomic analysis revealed distinct phosphorylation sites are modified by Cα targeted to Q1-YFP compared to free Cα. Thus, functional outcomes of synthetically recruited PKA on IKs regulation is critically dependent on the site of recruitment within the channel complex. The results reveal insights into divergent regulation of IKs by phosphorylation across different spatial and time scales, and suggest a TPP approach to develop new drugs to prevent exercise-induced sudden cardiac death.
Data availability
All data generated or analyzed during this study are included in the manuscript and supporting supplemental data file. Source data files have been provided.
Article and author information
Author details
Funding
National Heart, Lung, and Blood Institute (R01 HL142111)
- Henry M Colecraft
National Heart, Lung, and Blood Institute (R01 HL122421)
- Henry M Colecraft
National Institutes of Health (R01 GM109763)
- Robert S Kass
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2023, Zou et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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