PKC isoform-specific targeting of the phosphorylation site Serine191 in the GIRK4 subunit induces receptor-dependent modulation of GIRK channel activity

TL;DR

This study shows that different PKC isoforms target specific phosphorylation sites in GIRK4 channels, leading to receptor-specific regulation of channel activity.

Contribution

The study identifies that PKCε, activated by Angiotensin II, targets the S191 phosphorylation site in GIRK4, while PKCα activated by α1B-ARs does not.

Findings

Phosphorylation-deficient GIRK4 (S191A) channels show reduced inhibition by Ang II via PKCε.

Mutation of S191 does not affect α1B-AR-induced GIRK channel inhibition, indicating PKCα does not target this site.

Different GqPCR-activated PKC isoforms target distinct phosphorylation sites in GIRK4.

Abstract

G Protein activated inwardly rectifying K+ (GIRK) channels are inhibited during stimulation of Gq Protein-coupled receptors (GqPCRs) by depletion of phosphatidyl-4,5-bisphosphate (PIP2) and/or channel phosphorylation by proteinkinase C (PKC). Receptor-specific effects of Gq signaling pathways on GIRK channel activity comprise the activation of different PKC isoforms that target distinct phosphorylation sites within the GIRK4 subunit. The serine residue S191 in the GIRK4 subunit is an important phosphorylation site for PKC which contributes to GIRK channel inhibition. Until now, the specific PKC isoform that phosphorylates this residue is unknown. Furthermore, the functional impact of S191 for Gq-receptor-specific GIRK channel modulation has not been investigated. To evaluate the contribution of this PKC phosphorylation site to receptor-specific GIRK channel modulation, we monitored the activity of phosphorylation-deficient GIRK4 (S191A) channel mutants during stimulation of adrenergic α1B-receptors (α1B-ARs) or Angiotensin AT1-receptors. GqPCR-dependent modulation of GIRK currents was analyzed in voltage-clamp experiments in rat atrial myocytes and in CHO cells expressing phosphorylation-deficient GIRK channels of various subunit compositions. As a novel finding, we demonstrate that the inhibition of phosphorylation-deficient GIRK4 (S191A) channels is impaired during stimulation of AT1-Rs, indicating that this phosphorylation site is targeted at least by the Angiotensin II-activated PKCε isoform. In contrast, ablation of the S191 phosphorylation site does not attenuate phenylephrine-induced GIRK reduction, suggesting that S191 is not a primary target for the α1B-AR activated PKCα. Our data indicate that the different GqPCR-activated PKC isoforms target distinct phosphorylation sites within the GIRK4 subunit, resulting in receptor-specific regulation of the phosphorylation-deficient GIRK4 (S191A) channel mutant.

The online version contains supplementary material available at 10.1007/s00424-026-03157-0.

Phosphorylation-deficient GIRK4 (S191A) channels display reduced PMA-mediated inhibitionMutation of S191 alleviates the inhibitory effect of Ang II on GIRK channel activityResidue S191 is targeted by the AT1-R-activated PKCεGIRK4 (S191A) inhibition was not abrogated during stimulation of α1B-ARsα1B-AR-activated PKCα does not target S191

Phosphorylation-deficient GIRK4 (S191A) channels display reduced PMA-mediated inhibition

Mutation of S191 alleviates the inhibitory effect of Ang II on GIRK channel activity

Residue S191 is targeted by the AT1-R-activated PKCε

GIRK4 (S191A) inhibition was not abrogated during stimulation of α1B-ARs

α1B-AR-activated PKCα does not target S191

The online version contains supplementary material available at 10.1007/s00424-026-03157-0.

Figures