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. 2019 Feb 12;11(11):979–993. doi: 10.1093/jmcb/mjz008

Figure 1.

Figure 1

PS interaction is required for ADAM10 activation. (A and B) COS7 cells were transfected with the AP-tagged ADAM10 substrate BTC and stimulated with ionomycin (IO, 1 μM; A) or melittin (Mel, 0.5 μM; B) for 30 min. Shedding was dose-dependently reduced by addition of the competing phosphatidylserine head group (OPS) but not by the head group of phosphatidylcholine (OPC). OPS (10 mM), broadspectrum metalloprotease inhibitor TAPI-1 (10 μM), and ADAM10 inhibitor GI (3 μM) significantly abrogated the induced shedding. (C) COS7 cells were transfected with the AP-tagged ADAM10 substrate VE-cadherin and stimulated with ionomycin for 30 min in the presence of OPS (10 mM), OPC (10 mM), TAPI-1 (10 μM), or ADAM10 inhibitor GI (3 μM) and analysed for substrate shedding. (D) Shedding of full-length (FL) E-cadherin was monitored by immunoblot analysis. HaCaT keratinocytes were stimulated with ionomycin (1 μM) in the presence of TAPI-1 (10 μM), OPS (10 mM), OPC (10 mM), or ADAM10 inhibitor GI (3 μM). (E) Densitometric quantification of E-cadherin C-terminal fragment (CTF) generation of three independent western blots. * indicates a significant increase compared to unstimulated cells; # indicates significant decrease compared to stimulated control (P < 0.05, n = 3; ±SEM). NS, no significant difference. Data were analysed by one-way analysis of variance and Bonferroni multiple comparison post hoc test. CO, control.