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Receptor Species-dependent Desensitization Controls KCNQ1/KCNE1 K+ Channels as Downstream Effectors of Gq Protein-coupled Receptors.

ABSTRACT: Activation of G_q protein-coupled receptors (G_qPCRs) might induce divergent cellular responses, related to receptor-specific activation of different branches of the G_q signaling pathway. Receptor-specific desensitization provides a mechanism of effector modulation by restricting the spatiotemporal activation of signaling components downstream of G_q We quantified signaling events downstream of G_qPCR activation with FRET-based biosensors in CHO and HEK 293 cells. KCNQ1/KCNE1 channels (I_Ks) were measured as a functional readout of receptor-specific activation. Activation of muscarinic M₁ receptors (M₁-Rs) caused robust and reversible inhibition of I_Ks. In contrast, activation of ?_1B-adrenergic receptors (?_1B-ARs) induced transient inhibition of I_Ks, which turned into delayed facilitation after agonist withdrawal. As a novel finding, we demonstrate that G_qPCR-specific kinetics of I_Ks modulation are determined by receptor-specific desensitization, evident at the level of G?_q activation, phosphatidylinositol 4,5-bisphosphate (PIP₂) depletion, and diacylglycerol production. Sustained I_Ks inhibition during M₁-R stimulation is attributed to robust membrane PIP₂ depletion, whereas the rapid desensitization of ?_1B-AR delimits PIP₂ reduction and augments current activation by protein kinase C (PKC). Overexpression of Ca²⁺-independent PKC? did not affect the time course of ?_1B-AR-induced diacylglycerol formation, excluding a contribution of PKC? to ?_1B-AR desensitization. Pharmacological inhibition of Ca²⁺-dependent PKC isoforms abolished fast ?_1B receptor desensitization and augmented I_Ks reduction, but did not affect I_Ks facilitation. These data indicate a contribution of Ca²⁺-dependent PKCs to ?_1B-AR desensitization, whereas I_Ks facilitation is induced by Ca²⁺-independent PKC isoforms. In contrast, neither inhibition of Ca²⁺-dependent/Ca²⁺-independent isoforms nor overexpression of PKC? induced M₁ receptor desensitization, excluding a contribution of PKC to M₁-R-induced I_Ks modulation.

SUBMITTER: Kienitz MC

PROVIDER: S-EPMC5159502 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature

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Receptor Species-dependent Desensitization Controls KCNQ1/KCNE1 K+ Channels as Downstream Effectors of Gq Protein-coupled Receptors.

Kienitz Marie-Cécile MC Vladimirova Dilyana D Müller Christian C Pott Lutz L Rinne Andreas A

The Journal of biological chemistry 20161110 51

Activation of Gq protein-coupled receptors (GqPCRs) might induce divergent cellular responses, related to receptor-specific activation of different branches of the Gq signaling pathway. Receptor-specific desensitization provides a mechanism of effector modulation by restricting the spatiotemporal activation of signaling components downstream of Gq We quantified signaling events downstream of GqPCR activation with FRET-based biosensors in CHO ...[more]

PMID: 27834678

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Project description:The IKs channel complex is formed by the co-assembly of Kv7.1 (KCNQ1), a voltage-gated potassium channel, with its β-subunit, KCNE1 and the association of numerous accessory regulatory molecules such as PIP2, calmodulin, and yotiao. As a result, the IKs potassium current shows kinetic and regulatory flexibility, which not only allows IKs to fulfill physiological roles as disparate as cardiac repolarization and the maintenance of endolymph K+ homeostasis, but also to cause significant disease when it malfunctions. Here, we review new areas of understanding in the assembly, kinetics of activation and inactivation, voltage-sensor pore coupling, unitary events and regulation of this important ion channel complex, all of which have been given further impetus by the recent solution of cryo-EM structural representations of KCNQ1 alone and KCNQ1+KCNE3. Recently, the stoichiometric ratio of KCNE1 to KCNQ1 subunits has been confirmed to be variable up to a ratio of 4:4, rather than fixed at 2:4, and we will review the results and new methodologies that support this conclusion. Significant advances have been made in understanding differences between KCNQ1 and IKs gating using voltage clamp fluorimetry and mutational analysis to illuminate voltage sensor activation and inactivation, and the relationship between voltage sensor translation and pore domain opening. We now understand that the KCNQ1 pore can open with different permeabilities and conductance when the voltage sensor is in partially or fully activated positions, and the ability to make robust single channel recordings from IKs channels has also revealed the complicated pore subconductance architecture during these opening steps, during inactivation, and regulation by 1-4 associated KCNE1 subunits. Experiments placing mutations into individual voltage sensors to drastically change voltage dependence or prevent their movement altogether have demonstrated that the activation of KCNQ1 alone and IKs can best be explained using allosteric models of channel gating. Finally, we discuss how the intrinsic gating properties of KCNQ1 and IKs are highly modulated through the impact of intracellular signaling molecules and co-factors such as PIP2, protein kinase A, calmodulin and ATP, all of which modulate IKs current kinetics and contribute to diverse IKs channel complex function.

Dataset Information

Receptor Species-dependent Desensitization Controls KCNQ1/KCNE1 K+ Channels as Downstream Effectors of Gq Protein-coupled Receptors.

Publications

Receptor Species-dependent Desensitization Controls KCNQ1/KCNE1 K+ Channels as Downstream Effectors of Gq Protein-coupled Receptors.

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