GPR27 mediates L-lactate-induced Calcium and cAMP signalling in 3T3 cells

GPR27/SREB1 is a highly conserved orphan class A G-protein coupled receptor implicated in insulin production, metabolic regulation, tumour biology, neurodegeneration and L-lactate homeostasis, but its immediate second-messenger signalling remains poorly defined. We used single-cell Forster resonance energy transfer nanosensors to monitor cytosolic Ca2+ and cAMP in wild-type 3T3 MEF cells, CRISPR-Cas9 GPR27-knockout cells (GPR27KO) and GPR27-knockout cells transiently re-expressing FLAG-tagged GPR27 (GPR27-rescued). The GPR27 surrogate agonist 8535n (1 {micro}M) increased intracellular Ca2+ in wild-type and rescued cells but not in GPR27-knockout cells and produced no significant cAMP response in wild-type cells. Basal Ca2+ and cAMP levels were unaffected by GPR27 deletion. Extracellular L-lactate (2 mM) induced a GPR27-dependent increase in Ca2+ and cAMP in wild-type and rescued cells, but not in knockout cells, raising the possibility that L-lactate acts as an endogenous ligand or modulator of GPR27. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/743761v1_ufig1.gif" ALT="Figure 1"> View larger version (10K): org.highwire.dtl.DTLVardef@2af45dorg.highwire.dtl.DTLVardef@113e2corg.highwire.dtl.DTLVardef@8dea4org.highwire.dtl.DTLVardef@50ec49_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIGPR27 surrogate agonist 8535n increases intracellular Ca2+ but not cAMP in 3T3 cells. C_LIO_LIExtracellular L-lactate induces GPR27-dependent intracellular Ca2+ and cAMP increases in 3T3 cells. C_LIO_LIThese findings identify GPR27 as a putative candidate lactate sensor. C_LI