Cryo-EM analysis of PIP2 regulation in mammalian GIRK channels

Abstract
Data availability
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Abstract

G protein-gated inward rectifier potassium (GIRK) channels are regulated by G proteins and PIP₂. Here using cryo-EM single particle analysis we describe the equilibrium ensemble of structures of neuronal GIRK2 as a function of the C8-PIP₂ concentration. We find that PIP₂ shifts the equilibrium between two distinguishable structures of neuronal GIRK (GIRK2), extended and docked, towards the docked form. In the docked form the cytoplasmic domain, to which G_βγ binds, becomes accessible to the cytoplasmic membrane surface where G_βγ resides. Furthermore, PIP₂ binding reshapes the G_βγ binding surface on the cytoplasmic domain, preparing it to receive G_βγ. We find that cardiac GIRK (GIRK1/4) can also exist in both extended and docked conformations. These findings lead us to conclude that PIP₂ influences GIRK channels in a structurally similar manner to Kir2.2 channels. In Kir2.2 channels, the PIP₂-induced conformational changes open the pore. In GIRK channels, they prepare the channel for activation by G_βγ.

Data availability

The B-factor sharpened 3D cryo-EM density map and atomic coordinates of GIRK2 in the extended conformation (GIRK2Extended) and GIRK2 in the docked conformation with PIP2 (GIRK2Docked) have been deposited in the Worldwide Protein Data Bank (wwPDB) under accession number EMD-22199 and 6XIS, EMD-22200 and 6XIT, respectively. The B-factor sharpened 3D cryo-EM density map of GIRK1/4 in the extended conformation (GIRK1/4Extended) and docked conformation with PIP2 (GIRK1/4Docked) have been deposited in the Worldwide Protein Data Bank (wwPDB) under accession number EMD-22201 and EMD-22202, respectively.

The following data sets were generated

1. Niu Y
2. Tao X
3. MacKinnon R
(2020) Cryo-EM structure of the G protein-gated inward rectifier K+ channel GIRK2 (Kir3.2) in complex with PIP2
Protein Data Bank, 6XIT.

http://www.rcsb.org/structure/6XIT
1. Niu Y
2. Tao X
3. MacKinnon R
(2020) Cryo-EM structure of the G protein-gated inward rectifier K+ channel GIRK2 (Kir3.2) in complex with PIP2
The Electron Microscopy Data Bank, EMD-22200.

https://www.ebi.ac.uk/pdbe/entry/emdb/EMD-22200
1. Niu Y
2. Tao X
3. MacKinnon R
(2020) Cryo-EM structure of the G protein-gated inward rectifier K+ channel GIRK2 (Kir3.2) in apo form
Protein Data Bank, 6XIS.

http://www.rcsb.org/structure/6XIS
1. Niu Y
2. Tao X
3. MacKinnon R
(2020) Cryo-EM structure of the G protein-gated inward rectifier K+ channel GIRK2 (Kir3.2) in apo form
The Electron Microscopy Data Bank, EMD-22199.

https://www.ebi.ac.uk/pdbe/entry/emdb/EMD-22199
1. Niu Y
2. Tao X
3. MacKinnon R
(2020) Cryo-EM structure of the G protein-gated inward rectifier K+ channel GIRK1/4 (Kir3.1/Kir3.4) in apo form
The Electron Microscopy Data Bank, EMD-22201.

https://www.ebi.ac.uk/pdbe/entry/emdb/EMD-22201
1. Niu Y
2. Tao X
3. MacKinnon R
(2020) Cryo-EM structure of the G protein-gated inward rectifier K+ channel GIRK1/4 (Kir3.1/Kir3.4) in complex with bound PIP2
The Electron Microscopy Data Bank, EMD-22202.

https://www.ebi.ac.uk/pdbe/entry/emdb/EMD-22202

Article and author information

Author details

Yiming Niu

Laboratory of Molecular Neurobiology and Biophysics, Howard Hughes Medical Institute, The Rockefeller University, New York, United States

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-5683-1781
Xiao Tao

Laboratory of Molecular Neurobiology and Biophysics, Howard Hughes Medical Institute, The Rockefeller University, New York, United States

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-9381-7903
Kouki K Touhara

Laboratory of Molecular Neurobiology and Biophysics, Howard Hughes Medical Institute, The Rockefeller University, New York, United States

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0003-3167-9784
Roderick MacKinnon

Laboratory of Molecular Neurobiology and Biophysics, Howard Hughes Medical Institute, The Rockefeller University, New York, United States

For correspondence
mackinn@mail.rockefeller.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0001-7605-4679

Funding

National Institutes of Health (GM43949)

Roderick MacKinnon

Howard Hughes Medical Institute

Roderick MacKinnon

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

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.