Lysophospholipid stereoisomers exert distinct GPR55-mediated functions via different Gα subunits

Vertebrate glycerophospholipids typically exhibit a glycerol-3-phosphate (G3P)-configured backbone corresponding to the R-configured stereoisomer at the sn-2 chiral center. We previously found that the lysoglycerophospholipid lyso-phosphatidyl-β-d-glucoside (LysoPtdGlc) with G3P configuration, R-LysoPtdGlc, is an endogenous ligand of the G protein-coupled receptor GPR55, acting as an axon guidance cue via GPR55-Gα₁₃ signaling. However, LysoPtdGlc is hydrolytically derived from phosphatidyl-β-d-glucoside (PtdGlc), which exists in vivo in a 6:1 mixture of two stereoisomers, G3P-configured R-PtdGlc and glycerol-1-phosphate-configured S-PtdGlc. To test whether the stereoconfiguration of LysoPtdGlc influences its biological activity, we combined molecular dynamics simulations of GPR55 activation by R-LysoPtdGlc and S-LysoPtdGlc with in vitro and in vivo biological assays of GPR55-mediated functions in nervous system. Molecular dynamics simulations predicted that R-LysoPtdGlc, but not S-LysoPtdGlc, remained in the putative ligand-binding pocket of the GPR55-Gα₁₃ complex. Utilizing our previously established synthetic access to R-LysoPtdGlc and S-LysoPtdGlc, we investigated in vitro axonal chemotropic responses to these two stereoisomers. We observed R-LysoPtdGlc-mediated chemorepulsion corroborating our previous studies, and unexpectedly, S-LysoPtdGlc-induced chemoattraction via GPR55-Gα_S. Since these phenomena were observed in nociceptive neurons, we tested whether intrathecal administration of R-LysoPtdGlc or S-LysoPtdGlc induced a nociceptive phenotype in adult mice and found that R-LysoPtdGlc but not S-LysoPtdGlc increased behavioral sensitivity to mechanical stimuli, and that this response was dependent on GPR55. These data indicate that the stereoconfiguration of LysoPtdGlc determines its biological activity and suggest, at least in vitro, that LysoPtdGlc stereoisomers exert distinct GPR55-mediated functions via different Gα subunits.