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Reactive oxygen species (ROS) and advanced glycation end products (AGE)‐induced ERK5‐SUMOylation antagonizes anti‐inflammatory effect of shear stress in endothelial cells

Endothelial dysfunction is one of the major initiators of atherosclerosis, and diabetes is significantly involved in this process. Shear stress‐induced ERK5 activation has a critical role in regulating Kruppel‐like factor 2 (KLF2) expression as well as inhibiting adhesion molecule expression, which represents one of the anti‐inflammatory mechanisms. Here, we report that H2O2 and AGE negatively regulate ERK5 transcriptional activity and flow‐induced anti‐inflammatory actions through ERK5‐SUMOylation in endothelial cells. Point‐mutation analysis shows that ERK5 is covalently modified by SUMOylation at two conserved sites, Lys 6 and Lys 22. ERK5 transcriptional activity is inhibited by transfection of Ubc9 (E2 conjugase) or PIAS1 (E3 ligase). Compared with wild‐type ERK5, the SUMOylation defective (K6R/K22R) mutant shows more potent transcriptional activity of both ERK5 and MEF2. H2O2 and AGE induce endogenous ERK5‐SUMOylation in a time‐ and dose‐dependent manner, and K6R/K22R mutant, DN‐Ubc9, and siRNA‐PIAS1 reverse ERK5/MEF2 activity as well as promoter activity of KLF2. Finally PIAS1 knock down reverses the inhibitory effect of H2O2 in shear stress‐induced KLF2 and eNOS protein expression. These data clearly define SUMOylation‐dependent ERK5 transcriptional repression independent of kinase activity and could be one of the molecular mechanisms of diabetes‐mediated endothelial dysfunction.