Activated Notch receptor down-regulates Prox1 through Hey proteins. (A) Expression of Hey1 and Hey2 was determined by quantitative RT-PCR in LECs that were transduced with a control or NICD-expressing adenovirus for 48 hours. (B) Quantitative RT-PCR assay showing the repression of Prox1 mRNA in primary human LECs by adenoviral expression of NICD (AdNICD), Hey1 (AdHey1), Hey2 (AdHey2), Hey1 and Hey2 (AdHey1/AdHey2), and Hes1 (AdHes1) for 48 hours. (C) Expression of Prox1 and p27^Kip1 proteins was determined by Western blot analyses in primary LECs that were transduced with AdCTR, AdNICD, AdHey1, or AdHes1 for 48 hours. (D) The expression of Prox1 protein after cotransfection of LECs with a control (CTR) or NICD-expressing plasmid vector along with siRNA duplexes for the firefly luciferase (siLuci), Hey1 (siHey1), and Hey2 (siHey2) for 48 hours. Data are mean ± SD relative to the value of AdCTR.