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. Author manuscript; available in PMC: 2021 Mar 5.
Published in final edited form as: Cell. 2020 Mar 5;180(5):862–877.e22. doi: 10.1016/j.cell.2020.02.016

Fig. 5. PAGln Mediates Cellular Response Through G-Protein Coupled Receptor(s) and via ADRs.

Fig. 5.

(A) DMR dose response of PAGln, Norepi and Phe in MEG01 cells (n=4; max DMR responses after ligand addition).

(B) DMR response of PAGln (100 μM; left), Norepi (10 μM; middle) and collagen (10 μg/mL; right) in MEG01 cells pre-treated with the G-protein modulators pertussis toxin (PTX; 100 ng/mL), cholera toxin (CTX; 1 μg/mL), YM-254890 (0.5 μM) or SCH-202676 (1 μM) (n=5–10 as indicated).

(C-D) cAMP levels in (C) MEG01 cells and (D) washed human platelets pretreated with PAGln (100 μM; 5 min), in presence of PTX (100 ng/mL), CTX (1 μg/mL), YM-254890 (1 μM) or SCH-202676 (1 μM). cAMP levels were normalized to 100% immediately before addition of PAGln (n=4–9 as indicated).

(E) Structure similarity between PAGln and catecholamines (ISO, Epinephrine and Norepi).

(F) DMR response in MEG01 cells transfected with control scrambled siRNAs, and siRNAs against the α2A, α2B and β2 ADRs and analyzed under indicated conditions (n=6–9 as indicated). Maximum DMR response to PAGln was normalized to 100%.

(G) PAGln (100 μM) DMR response quantified in MEG01 cells treated with 10 μM selective β2 antagonist ICI118,551, nonselective β-blocker propranolol or nonselective α2 antagonist RX821002 for 30 min (n=3–8 as indicated). The maximum DMR response to PAGln was normalized to 100%.

(H) cAMP levels in MEG01 cells after PAGln (100 μM; 5 min) treatment in the presence of 10 μM ICI118,551, propranolol or RX821002 (n=6–10 as indicated). cAMP levels were normalized to 100% in all treatments immediately prior to addition of PAGln.

Nonparametric-Mann Whitney test was used for non-pairwise comparisons and Kruskal-Wallis (K.W.) test for multiple comparisons. Data points represent the mean ± SEM (n=biological replicates). See also Figure S4–6.