DOMINANT SIGNALS LEADING TO INHIBITOR KAPPA-B PROTEIN-DEGRADATION MEDIATE CD40 LIGAND RESCUE OF WEHI-231 IMMATURE B-CELLS FROM RECEPTOR-MEDIATED APOPTOSIS

Recently, we demonstrated maintenance of nuclear factor (NF)-kappa B/R el factors plays a major role in B cell survival. Treatment of WEHI 23 1 immature B cells with an Ab against the surface IgM protein (anti-Ig M) induces apoptosis that can be rescued by engagement of CD40 recepto r. The dramatic decrease in high basal levels of NF-kappa B/Rel activi ty induced by anti-IgM treatment led to cell death, CD40 ligand (CD40L ) treatment prevented the drop in NF-kappa B/Rel factor binding by ind ucing a sustained decrease in inhibitor (I) kappa B-(alpha and transie nt decrease in I kappa B-beta protein levels. In this study, we have i nvestigated the regulation of these NF-kappa B/Rel-inhibitory proteins . In exponentially growing WEHI 231 cells, the I kappa B-(alpha and I kappa B-beta proteins decayed with an approximate t1/2 of 38 and 76 mi n, respectively, which was blocked effectively upon addition of the pr oteasome-specific inhibitor (benzylcarbonyl)-Leu-Leu-phenylalaninal (Z -LLF-CHO). Anti-IgM treatment stabilized I kappa B-(alpha and I kappa B-beta proteins, CD40L treatment resulted in a dramatic decrease in t1 /2 (<5 min) for both I kappa B molecules, which was inhibited by addit ion of Z-LLF-CHO, CD40L treatment also caused a delayed increase in I kappa B-beta mRNA levels, most likely contributing to the observed rec overy of I kappa B-beta levels. Microinjection of I kappa B-alpha-glut athione S-transferase fusion protein into nuclei of WEHI 231 cells abl ated protection by CD40L from receptor-mediated killing. Furthermore, CD40L rescued apoptosis induced upon microinjection of a vector expres sing wild-type I kappa B-alpha, but not a 32A/36A mutant form of I kap pa B-alpha, unable to be phosphorylated and hence degraded, Thus, cont rol of turnover of I kappa B proteins by CD40L plays a major role in m aintenance of NF-kappa B/Rel and resultant rescue of WEHI 231 cells fr om apoptosis.