Figure 4
CnAα/β attenuate NIK-dependent Spi-B expression and nuclear translocation of NF-κBs.
A. Quantitative RT-PCR analysis of Spi-B1 expression in aly/+ and aly/aly MEFs treated with an agonistic anti-LtβR antibody. Aly/+ and aly/aly MEFs or aly/+ and aly/aly MEFs depleted of both CnAα/β by siRNAs were stimulated with the agonistic anti-LtβR antibody. Expression of Spi-B1 was evaluated by qPCR analysis. Representative data of three independent triplicate wells are shown. Black bars indicate mean values. P indicates the results of Student’s t-tests. B. qPCR analysis of Spi-B expression in MEFs depleted of CnAα, CnAβ, or both CnAα/β. Wild-type MEFs depleted of CnAα and/or CnAβ by siRNAs (upper panels) were stimulated with the agonistic anti-LtβR antibody. Representative data of three independent triplicate wells are shown. Black bars indicate mean values. P indicates the results of Student’s t-tests. Blots are cropped for clarity. Full-length blots are presented in Supplementary Figure 2. C. Depletion of CnAs enhances nuclear localization of RelA and RelB induced by LtβR signaling. CnAα-, CnAβ-, or CnAα/β-depleted MEFs were treated with the agonistic anti-LtβR antibody for 3 and 24 h (LtβR-Ab), or untreated (control). Nuclear and cytosolic protein fractions were analyzed by western blotting. Band intensities of RelA and RelB relative to PARP were normalized to that of control (control siRNA-treated cells without stimulation) and exhibited on the top of panels. Antibodies used for western blotting are indicated at the left of panels. Blots are cropped for clarity. Full-length blots are presented in Supplementary Figure 2.
