Pkc alpha phosphorylates cytosolic nf-kappab/p65 and pkc delta delays nuclear translocation of nf-kappab/p65 in u1242 glioblastoma cells

Abstract

Aim: Protein kinase-C (PKC) and NF-kappaB are involved in cell survival, proliferation, migration and radioresistance in glioblastoma multiforme (GBM). We sought to determine the interaction between PKC and NF-kappaB pathways. Material and Methods: The activation of NF-kappaB by PKC α and PKC δ was assessed by Western blotting after the stimulation with Phorbol 12-Myristate 13-Acetate (PMA). Gene silencing of PKC α, PKC δ and NFkappaB/ p65 with siRNA interference was utilized to evaluate their roles in NFkB activation and cell proliferation. Results: PMA induced the phosphorylation of NF-kappaB/p65 by PKC α. Gene silencing with siRNA against NF-kappaB/p65 inhibited [3H]-thymidine incorporation in U1242 GBM cells. PKC δ decelerated the nuclear translocation of activated NF-kappaB/p65 up to 4 hours after the stimulation. PMA induced death was not observed in PKC δ silenced cells where activated NF-kappaB/p65 was located immediately in the nucleus. Conclusion: NF-kappaB/p65 is pro-survival and proliferative factor in U1242 GBM cells. PKC α is needed to phosphorylate NF-kappaB/pPKC δ delays the translocation of active NF-kappaB/p65 into the nucleus. PMAinduced cell death occurred if the phospho-NF-kappaB/p65 was prohibited from entering the nucleus in PKC δ positive cells. Translocation of phosphorylated form of NF-kappaB into the nucleus is critical in GBM cell proliferation.