Extended Data Fig. 7: The activation of cellular signaling by L-LEN and GPR50 and the functional conservation of L-LEN across species.
From: Photo-cross-linking-assisted deorphanization deciphers GPR50–L-LEN pairing in metabolism
Related to Fig. 4. (a) Schematic (left), representative fluorescence response of the G-Flamp2 (middle) and the quantification of the fluorescence response (right) in GPR50-expressing cells to the treatment of L-LEN variants and B-LEN (n = 15 cells; p = 0.99/1/1/1/0.99/1/1.39×10−38 between buffer and L-LEN/B-LEN/A/D/F/G/FSK with One-way ANOVA followed by Tukey HSD post hoc test). (b) The intensity of L-LEN and related peptides in mouse brain extracts detected by mass spectrometry (n = 3 mice). (c) The relative intensities of L-LEN and related peptides over GAPDH in human cerebrospinal fluid (CSF) (n = 2 samples). (d) Sequences of L-LEN in different mammalian species. The amino acids differing from human L-LEN are highlighted in red. (e) Left, traces showing the forskolin (FSK) and IBMX evoked G-Flamp2 responses in GPR50-expressing cells pretreated with buffer, human L-LEN or mouse L-LEN (both at 10−6 M). Right, the group analysis of the cAMP inhibition by human and mouse L-LEN (right) (n = 3 independent cultures; p = 0.066). (f) Left, partial confidence metrics of the predicted structure (left) of L-LEN and GPR50. Right, the Predicted Align Error (PAE) matrix of all amino acid residue pairs in the docking structure, with asterisks indicating potential contacting residues. The pLDDT score curves for all the atoms in each chain are shown on the right. Data are shown as the mean ± SEM, n.s. not significant; ***, p < 0.001. See Supplementary Table 1 for statistics.
