Polyunsaturated fatty acids inhibit a pentameric ligand-gated ion channel through one of two binding sites

Abstract

Polyunsaturated fatty acids (PUFAs) inhibit pentameric ligand-gated ion channels (pLGICs) but the mechanism of inhibition is not well understood. The PUFA, docosahexaenoic acid (DHA), inhibits agonist responses of the pLGIC, ELIC, more effectively than palmitic acid, similar to the effects observed in the GABAA receptor and nicotinic acetylcholine receptor. Using photo-affinity labeling and coarse-grained molecular dynamics simulations, we identified two fatty acid binding sites in the outer transmembrane domain (TMD) of ELIC. Fatty acid binding to the photolabeled sites is selective for DHA over palmitic acid, and specific for an agonist-bound state. Hexadecyl-methanethiosulfonate modification of one of the two fatty acid binding sites in the outer TMD recapitulates the inhibitory effect of PUFAs in ELIC. The results demonstrate that DHA selectively binds to multiple sites in the outer TMD of ELIC, but that state-dependent binding to a single intrasubunit site mediates DHA inhibition of ELIC.

Structural mechanism of leaflet-specific phospholipid modulation of a pentameric ligand-gated ion channel

“…and C300S (previously shown to slow current decay in the presence of agonist 12,31 ), and produced P254G/C300S/V261Y/G319F/I320F (ELIC5) (Extended Data Fig. 4a).…”

“…pLGIC function is modulated by certain lipids 6 , including anionic phospholipids which enhance agonist response in the nAChR [7][8][9][10][11] . Anionic phospholipids, as well as sterols and fatty acids, have also been shown to modulate other pLGICs [12][13][14][15][16] . It is intuitive that lipids modulate pLGICs by influencing the stability of different states that the channel samples during gating; however, mechanisms of phospholipid modulation of any pLGIC remain poorly understood.…”

“…With the recent boon in lipid-bound pLGIC structures by cryo-EM, investigation into the structural mechanisms of lipid modulation has focused on direct binding of lipids to specific sites 12,13,15,17,18 . Structures of the 5-HT 3A receptor (5-HT 3A R), GABA A receptor, and the prototypic pLGIC, Erwinia chrysanthemi ligand-gated ion channel (ELIC), show bound phospholipids at various sites 14,[19][20][21][22][23] .…”

Recent Insight into Lipid Binding and Lipid Modulation of Pentameric Ligand-Gated Ion Channels

“…ELIC desensitization is slowed even further in the presence of the anionic lipid, PG (see below) [ 32 , 111 , 113 ]. The fatty acid DHA increases the rates of GLIC desensitization [ 114 ], while treatment of ELIC with DHA leads to a reduction in the magnitude of the agonist-induced peak current, but does not alter the EC 50 for channel gating [ 115 ].…”

“…Mutations in several of the cardiolipin binding site residues influence the gating of ELIC. Polyunsaturated fatty acids have also been detected bound to a site at the M4–M1 interface, adjacent to where DHA binds to GLIC [ 115 ].…”