NFκB activity and Notch-1 expression regulate GATA3 expression. (A) TaqMan PCR of 1 typical result of L-428, L-1236, KM-H2, and U-HO1 cells 5 days after viral transduction with IκBα superrepressor, GFP+ cell sorting, and RNA/complimentary DNA preparation. The 3 typical NKκB target Birc3, ICAM-1, and Lta were used as controls for NFκB activity. mRNA expression for all 4 analyzed genes is depicted as fold change down-regulation after normalization to actin housekeeping controls. (B) Immunoblot for Notch-1 expression of HDLM-2, KM-H2, L-428, L-1236, and U-HO1 cells. Jurkat cells served as positive controls. 100 μg protein were loaded. Actin was used as a loading control. (C) TaqMan RT-PCR as in (A) on KM-H2, L-428, and U-HO1 cells 5 days after delivery of small hairpin RNA against Notch-1. (D) Immunoblot for Notch-1 and GATA3 expression in L-428 and U-HO1 cells 5 days after transduction of indicated small hairpin RNA sequences. Actin served as loading control. Each lentiviral transduction was performed 3 times.