Bcl-3 in ALCL and HRS cells. (A) mRNA and protein expression of Bcl-3 in Hodgkin, ALCL, and other non-Hodgkin cell lines, as indicated. Top panel: mRNA analysis of BCL3 and, as control, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) by Northern blot (NB). Bottom panel: protein expression of Bcl-3 in various lymphoma cell lines, as indicated. As a control, expression of α-tubulin is shown. WB indicates Western blot. (B) Localization of Bcl-3. Cytoplasmic (C) and nuclear (N) extracts of Hodgkin (L428, L1236) and ALCL (K299, SU-DHL-1) cells were analyzed for localization of Bcl-3 and IκBα by Western blot analysis. Note that L428 HRS cells lack expression of IκBα WT protein. (C) Bcl-3 IP-shift assay. Bcl-3 containing NF-κB complexes of HRS (HDLM-2, L1236), ALCL (K299, SU-DHL-1), and other non-Hodgkin cell lines (Reh, Namalwa, Daudi) were immunoprecipitated by use of an anti–Bcl-3 antibody. Thereafter, precipitated complexes were analyzed with or without supershift (ss) analysis for NF-κB DNA binding activity by EMSA. Nuclear extracts of TNF-α–stimulated Hela cells are shown as control. Positions of NF-κB and (p50)2 complexes are indicated. n.s. indicates nonspecific; IP, immunoprecipitation.
