Structural insights into GIRK2 channel modulation by cholesterol and PIP2

Abstract

SUMMARY G-protein-gated inwardly rectifying potassium (GIRK) channels are important for determining neuronal excitability. In addition to G proteins, GIRK channels are potentiated by membrane cholesterol, which is elevated in the brains of people with neurodegenerative diseases such as Alzheimer’s dementia and Parkinson’s disease. The structural mechanism of cholesterol modulation of GIRK channels is not well understood. In this study, we present cryo-electron microscopy (cryoEM) structures of GIRK2 in the presence and absence of the cholesterol analog cholesteryl hemisuccinate (CHS) and phosphatidylinositol 4,5-bisphosphate (PIP 2 ). The structures reveal that CHS binds near PIP 2 in lipid-facing hydrophobic pockets of the transmembrane domain. Our structural analysis suggests that CHS stabilizes PIP 2 interaction with the channel and promotes engagement of the cytoplasmic domain onto the transmembrane region. Mutagenesis of one of the CHS binding pockets eliminates cholesterol-dependent potentiation of GIRK2. Elucidating the structural mechanisms underlying cholesterol modulation of GIRK2 channels could facilitate the development of therapeutics for treating neurological diseases.

Lipid accumulation induced by APOE4 impairs microglial surveillance of neuronal-network activity

“…Notably, GIRK channels are known to be lipid-gated as K+ flux is dependent on binding of the phospholipid phosphatidylinositol (4,5)P2 (PIP2) (Huang et al, 1998). In neurons, cholesterol has been shown to modulate GIRK currents, with cholesterol enrichment enhancing channel activity and thus decreasing neuronal excitability (Bukiya et al, 2019;Bukiya et al, 2017;Mathiharan et al, 2021). Based on our RNA-seq analysis, APOE4 iMGL conditioned media significantly upregulated transcript levels of GIRK3 in spheroids, while GIRK1, GIRK2 and GIRK4 remained unchanged (Figure 5I).…”

Entropy in the Molecular Recognition of Membrane Protein–Lipid Interactions

“…Although first described for Kir2.2 (Figure S16), recent structures of Kir3.2 have shown it also populates two distinct conformations where the CTD is docked, forming contacts with the cytoplasmic face of the TMD, or in an extended state, CTD displaced from the transmembrane domain . For some of the PIPs, a lipid binding model where the lipid binds to two nonidentical sites on the channel resulted in substantially improved fits.…”

“…It is challenging from the lipid binding data here to determine if Kir3.2 follows the conformational selection or induced-fit binding pathway. However, recent structural studies show a population of Kir3.2 in docked and extended states (pre-existing populations) and in the presence of increasing concentrations of PI­(4,5)­P 2 a population shift to the docked state …”

Inwardly Rectifying Potassium Channel Kir2.1 and its “Kir-ious” Regulation by Protein Trafficking and Roles in Development and Disease

“…The putative cholesterol binding sites overlap partially with an inverted cholesterol recognition amino acid consensus, or CARC, motif [consensus sequence (R/K)X 1-5 (Y/F)X 1-5 (L/V) where X is any amino acid and corresponds to residues R67-F73-V77 in Kir2.1], however a second putative CARC motif (R82-F88-L90) does not appear to bind cholesterol ( Rosenhouse-Dantsker et al, 2013 ). Interestingly, using similar approaches, putative cholesterol binding sites were defined in Kir3.x family members, and some of these have been substantiated by recent cryoEM studies ( Bukiya et al, 2017 ; Bukiya and Rosenhouse-Dantsker, 2017 ; Mathiharan et al, 2021 ). While the cholesterol binding sites map to similar regions in the TMDs of Kir2.1 and Kir3.x family members (compared in ( Rosenhouse-Dantsker, 2019 )), cholesterol inhibits Kir2.1 but activates Kir3.x channels, presenting an interesting functional dichotomy between these two family members.…”