Constitutive BLyS expression through the NF-κB pathway activates a positive feedback loop contributing to NHL-B cell survival. (A) EMSA analysis of NF-κB protein binding to the BLYS promoter. Nuclear extracts from NHL-B cells were transfected with dominant-negative (DN) NF-κB or control plasmid and analyzed with an oligonucleotide probe for the BLYS–NF-κB binding site. (B) Immunoblot analysis of BLyS expression in NHL-B (LBCL-MS) cells transfected with DN NF-κB or control plasmid. Actin was used as loading control. BLyS levels were normalized to those of actin as previously described and presented as relative fold decrease compared to control samples. (C) EMSA analysis of NF-κB protein binding to the BLYS promoter in NHL-B (MCL-SP53) cells treated with various doses of the IκB inhibitor BAY11-7082. Lamin B was used as nuclear protein loading control. (D) Immunoblot analysis of BLyS in NHL-B (MCL-SP53) cells treated with various doses of IκB inhibitor BAY11-7082. (E) Immunoblot analysis for BLyS, p52, and Rel-B expression in NHL-B (LBCL-MS) cells treated with 100 nM p52 or Rel-B siRNA. Actin was used as loading control. (F) EMSA analysis of NF-κB binding to the BLYS promoter. Nuclear extracts from an NHL-B cell line (LBCL-MS) were transfected with BLYS or control siRNA and analyzed with an oligonucleotide probe for the BLYS–NF-κB binding site. Lamin B was used as nuclear protein loading control. (G) Immunoblot analysis of phosphorylated IκBα (pIκBα) and p52 in NHL-B cells transfected with BLYS or control siRNA. pIκBα and p52 protein expression levels were normalized to those of actin as previously described in panel B.