CpG stimulation of primary mouse B cells is blocked by inhibitory oligodeoxyribonucleotides at a site proximal to NF-kappa B activation

Bacterial DNA and CpG-oligodeoxyribonucleotides (ODN) are powerful B cell a ctivators, inducing apoptosis protection, cell cycle entry, proliferation, costimulatory molecule expression, immunoglobulin (Ig) and interleukin-6 (I L-6) secretion. However, proximal events in B cell activation by ODN are on ly partially characterized, including the translocation of NF-kappaB to the nucleus. In this paper, we provide evidence that CpG-ODN-induced cell cycl e entry and apoptosis protection are blocked by SN50 or gliotoxin and thus require NF-kappaB activation. NF-kappaB activation occurred within 30 minut es of stimulation of murine B cells with a phosphorothioate (S) CpG-ODN and persisted for up to 40 hours, with p50, p65, and c-Rel as the major compon ents. Similar to other NF-kappaB inducers, CpG-ODN caused an early I kappaB alpha and I kappaB beta degradation plus cleavage of the p50 precursor and subsequent NF-kappaB nuclear translocation. A group of closely related S-O DN, which specifically blocked CpG-induced B cell activation at submicromol ar concentrations, also prevented NF-kappaB DNA binding and transcriptional activation. These inhibitory S-ODN differed from stimulatory S-ODN by havi ng 2-3 G substitutions in the central motif. As inhibitory S-ODN did not di rectly interfere with the NF-kappaB DNA binding but prevented CpG-induced N F-kappaB nuclear translocation of p50, p65, and c-Rel and blocked p105, I k appaB alpha, and I kappaB beta degradation, we concluded that their putativ e target must lie upstream of inhibitory kinase (IKK) activation.