CD40-MEDIATED SIGNALS INHIBIT THE BINDING OF TNF RECEPTOR-ASSOCIATED FACTOR-2 TO THE CD40 CYTOPLASMIC DOMAIN

TNF receptor-associated factor 2 (TRAF2) is a signal-transducing prote in associated with the CD40 cytoplasmic domain, It has been hypothesiz ed that during signal transduction, TRAF2 must be released from CD40 i n order for it to interact with downstream signaling molecules. We fou nd that CD40 and TRAF2 were constitutively associated with each other in a human B cell line. Following stimulation with an anti-CD40 Ab, a decrease in the amount of CD40-associated TRAF2 was observed that coul d not be explained by a change in total level of either of the protein s, These results, as well as similar findings obtained with 293 cells overexpressing CD40 and TRAF2, suggested that CD40-mediated signals in hibited the CD40-TRAF2 interaction. We then conducted binding studies using CD40 cytoplasmic domain fusion proteins and TRAF2 derived from e ither control or CD40-stimulated cell lines, These in vitro studies al so indicated that tbe binding of TRAF2 to the CD40 cytoplasmic domain was inhibited by CD40 stimulation. The results of these experiments, a s well as differences between the in vitro and in vivo findings, indic ated that multiple mechanisms were involved in the inhibition of the C D40-TRAF2 interaction by CD40 signals. Possible mechanisms oi inhibiti on are discussed based on mapping of the TRAF2 binding site on the CD4 0 cytoplasmic domain.