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. 2013 Feb 1;24(3):398–408. doi: 10.1091/mbc.E12-07-0513

FIGURE 5:

FIGURE 5:

NO/PKG inhibitors blunt PAK2 Ser-20 phosphorylation and rescue PAK activation. (A) BAECs were left untreated (NT) or treated with CPTIO (200 μM, 30 min), ODQ (10 μM, 30 min), or KT5823 (2 μM, 1 h). Cells were sheared for 15 min, and Ser-20 phosphorylation was determined. n = 4–7. (B) Endothelial cells were treated as in A, and PAK2 Ser-141 phosphorylation was determined. n = 3. (C) Endothelial cells were transfected with eNOS siRNA, plated on BM, and exposed to shear stress for 15 min. Ser-20 phosphorylation was determined as previously described. n = 3. (D) Endothelial cells treated with KT5823 as in A were sheared for 15 min and PAK2 membrane translocation determined. Results are normalized to α5 integrin levels in the membrane fraction. Representative immunoblots are shown. n = 3. (E) Endothelial cells on BM were treated with KT5823 as in A and exposed to shear stress for 15 min. PKA activity was determined by affinity precipitation and normalized to total PKA in cell lysates. Values are means ± SE, n = 3. *p < 0.05.