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| Item Type: | Article |
|---|---|
| Title: | Live-cell quantitative monitoring reveals distinct, high-affinity Gβγ regulations of GIRK2 and GIRK1/2 channels |
| Creators: |
Handklo-Jamal, Reem  ORCID: https://orcid.org/0000-0001-6237-5412, Raifman, Tal Keren, Shalomov, Boris ORCID: https://orcid.org/0000-0002-1322-4906, Hofer, Patrick ORCID: https://orcid.org/0009-0004-5778-9599, Kahanovitch, Uri, Friesacher, Theres ORCID: https://orcid.org/0000-0001-5632-3472, Tabak, Galit, Tsemakhovich, Vladimir, Reddy, Haritha P., Chomsky-Hecht, Orna, Tripathy, Debi Ranjan, Zuhlke, Kerstin, Dessauer, Carmen W., Klussmann, Enno ORCID: https://orcid.org/0000-0003-4004-5003, Haitin, Yoni ORCID: https://orcid.org/0000-0002-1648-6792, Hirsch, Joel A. ORCID: https://orcid.org/0000-0001-7544-8668, Stary-Weinzinger, Anna ORCID: https://orcid.org/0000-0002-9202-0484, Yakubovich, Daniel ORCID: https://orcid.org/0000-0003-2871-1026 and Dascal, Nathan ORCID: https://orcid.org/0000-0002-5397-4146
|
| Abstract: | G(i/o) protein-coupled receptors (GPCRs) inhibit cardiac and neuronal excitability via G protein-activated K+ channels (GIRK), assembled by combinations of GIRK1 - GIRK4 subunits. GIRKs are activated by direct binding of the Gβγ dimer of inhibitory G(i/o) proteins. However, key aspects of this textbook signaling pathway remain debated. Recent studies suggested no G(i/o)-GIRK pre-coupling and low (>250 µM) Gβγ-GIRK interaction affinity, contradicting earlier sub-µM estimates and implying low signaling efficiency. We show that Gγ prenylation, which mediates Gβγ membrane attachment required for GIRK activation, also contributes to the Gβγ-GIRK interaction, explaining the poor affinity obtained with non-prenylated Gβγ. Using quantitative protein titration and electrophysiology in live Xenopus oocytes, Gβγ affinity for homotetrameric GIRK2 ranges from 4-30 µM. Heterotetrameric GIRK1/2 shows a higher Gβγ apparent affinity due to the Gβγ-docking site (anchor) in GIRK1, which enriches Gβγ at the channel. Biochemical approaches and molecular dynamic simulations reveal that the Gβγ anchor is formed by interacting N-terminal and distal C-terminal domains of the GIRK1 subunits, distinct from the Gβγ-binding “activation” site(s) underlying channel opening. Thus, the affinity of Gβγ-GIRK interaction is within the expected physiological range, while dynamic pre-coupling of Gβγ to GIRK1-containing channels through high-affinity interactions further enhances the GPCR-G(i/o)-GIRK signaling efficiency. |
| Keywords: | G Protein-Coupled Inwardly-Rectifying Potassium Channels, GTP-Binding Protein Beta Subunits, GTP-Binding Protein Gamma Subunits, HEK293 Cells, Oocytes, Protein Binding, Signal Transduction, Animals, Xenopus Laevis |
| Source: | Nature Communications |
| ISSN: | 2041-1723 |
| Publisher: | Nature Publishing Group |
| Volume: | 16 |
| Number: | 1 |
| Page Range: | 11607 |
| Date: | 29 December 2025 |
| Official Publication: | https://doi.org/10.1038/s41467-025-66730-8 |
| PubMed: | View item in PubMed |
| Related to: |
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