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. 2015 Sep 28;210(7):1117–1131. doi: 10.1083/jcb.201504007

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© 2015 Grimsey et al.

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Figure 3. — Activated PAR1 ubiquitination is mediated by NEDD4-2. (A) PAR1 WT or untransfected (UT) HeLa cells were transfected with ns, NEDD4-1 (N4-1), or NEDD4-2 (N4-2) siRNA and stimulated with 100 µM SFLLRN. Then PAR1 ubiquitination was detected. See Fig. S2 (A–C). (B) Quantitation of PAR1 ubiquitination. The data (mean ± SD [error bars], n = 3) were analyzed using a Student’s t test (*, P < 0.05). (C) PAR1 surface expression and internalization detected in HeLa cells transfected with ns, N4-1, and N4-2 siRNA after 10 min of stimulation with 100 µM SFLLRN. The data (mean ± SD [error bars], n = 3) were analyzed using a Student’s t test. (D) PAR1 ubiquitination detected in endothelial cells transfected with ns or N4-2 siRNAs after stimulation with 10 nM α-Th. (E) PAR1 ubiquitination was quantitated. The data (mean ± SD [error bars], n = 3) were analyzed using a Student’s t test (*, P < 0.05). (F) PAR1 surface expression and internalization determined in endothelial cells transfected with ns or N4-2 siRNA after 5 min of 10 nM α-Th. The data (mean ± SD [error bars], n = 3) were analyzed using a Student’s t test.