CHARACTERIZATION OF CD40 SIGNALING DETERMINANTS REGULATING NUCLEAR FACTOR-KAPPA-B ACTIVATION IN B-LYMPHOCYTES

CD40 signaling to B cells is important for generating an effective hum oral immune response. CD40 ligation leads to B cell activation events such as proliferation, Ig secretion, isotype switching, and up-regulat ion of cell surface molecules, as well as the generation of memory B c ells. Many of these events are dependent upon the ability of CD40 to a ctivate the transcription factor NF-kappa B (NF-kappa B).(3) To define the CD40 signaling components upstream of NF-kappa B activation and t he functional consequences downstream of NF-kappa B activation, we exa mined mouse B cell transfectants expressing wild-type or mutant human CD40. Analysis of CD40 cytoplasmic domain truncation and point mutants defined a 10-amino acid CD40 cytoplasmic signaling determinant requir ed for NF-kappa B activation. A threonine residue at position 234, pre viously shown to be important for CD40 association with TNF receptor-a ssociated factor 2 (TRAF2), TRAF3, and TRAF5, was not required for NF- kappa B activation. This suggests that in B cells, CD40-induced NF-kap pa B activation can occur independently of TRAF2 and TRAF5 association . NF-kappa B activation was independent of the transmembrane domain of CD40, suggesting that it is independent of p23, a molecule that assoc iates with CD40 in a region other than the cytoplasmic domain. Proteas ome-dependent inhibitory kappa B alpha (I kappa B alpha) and I kappa B beta degradation occurred downstream of CD40 ligation and preceded CD 40-mediated NF-kappa B nuclear translocation. CD40- or pervanadate-med iated I kappa B tyrosine phosphorylation was not detected, NF-kappa B activation correlated with the ability of CD40 to induce Ab secretion and the up-regulation of ICAM-1 and LFA-1. However, NF-kappa B activat ion was insufficient for CD40-mediated up-regulation of B7-1, Fas, and CD23.