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. Author manuscript; available in PMC: 2022 Jan 3.
Published in final edited form as: Sci Signal. 2021 Apr 20;14(679):eabc0425. doi: 10.1126/scisignal.abc0425

Figure 3. CLICs function in endothelial S1P pathway to regulate Rac1 activity.

Figure 3.

(A) HUVECs were serum starved for 3hrs and lysates were collected following 1μM S1P stimulation for 5min. Rac1 activity was measured by G-LISA assay and normalized to unstimulated cells for each condition (N=3, **p<0.01). (B) HUVECs were treated with IBMX and Forskolin followed by addition of 1μM S1P for 30min. Lysates were collected with 0.1M HCl, and cAMP levels were measured using a competitive immunoassay. (N=3, **p<0.01) (C and D) Following serum starvation and S1P treatment (1μM, 5min), (C) Ras activity was measured by G-LISA assay and normalized to unstimulated cells (N=3, *p<0.05). (D-E) Western blotting for ERK phosphorylation and quantification by densitometry analysis, normalized to the signal of total ERK protein respectively (N=3, **p<0.01). All significance (A-E) was determined by ANOVA with Bonferroni correction for multiple comparisons test.