VEGF signals through cyclin E/Cdk2 to promote mis-regulation of centrosome duplication in endothelial cells. (A-C) HUVECs were serum starved and then treated for 12 hours with low or high VEGF. (A) Western blot of lysates hybridized with anticyclin E, normalized to actin. (B) Autoradiogram of Cdk2 immune-complex kinase assay normalized to total Cdk2 immunoprecipitation (IP). (C) Western blot of lysates hybridized with anti-P-NPM, normalized to total NPM. (D-E) HUVECs were infected with empty vector (EV) or cyclin E shRNA (CycE KD) lentivirus for 6 hours and then incubated in low or high VEGF for 96 hours. (D) Western blot of lysates hybridized to anticyclin E and normalized to actin. (E) After lentiviral infection and VEGF treatment, cells were fixed, stained for γ-tubulin and DRAQ5, and centrosome numbers were counted (low VEGF, n = 656; high VEGF, n = 750; high VEGF + EV, n = 717; high VEGF + cyclin E knockdown [CycE KD], n = 696). All experiments were performed at least 3 times. ***P < .0001 vs low VEGF. #P < .05 vs high VEGF.