NF-κB is involved in the survival of cerebellar granule neurons:: association of Iκβ phosphorylation with cell survival

The NF-kappa KB transcription factor consists of dimeric complexes belonging to the Rel family, which include p50, p52, p65 (RelA), RelB and c-Rel. NF-kappaB activity is tightly controlled by I kappaB proteins which bind to NF-kappaB preventing its translocation to the nucleus. Activation of NF-kappaB is most often mediated by I kappaB degradation, which permits NF-kappaB to enter the nucleus. We investigated the role of NF-kappaB in the survival of cerebellar granule neurons. We found that survival of these neurons in high potassium medium is blocked by three separate inhibitors of NF-kappaB activity: SN-50, N-tosyl-L-phenylalanine chloromethyl ketone and pyrrolidinedithiocarbamate, indicating that NF-kappaB is required for neuronal survival. Gel-shift assays reveal three complexes that bind to the NF-kappaB binding site in high potassium medium. Switching these cultures to low potassium medium, a stimulus that leads to apoptotic death, causes a reduction in the level of the largest complex, which contains p65. Overexpression of p65 by transfection inhibits low potassium-induced apoptosis, whereas overexpression of I kappaB alpha promotes apoptosis even in high potassium medium. Surprisingly, however, neither the level of endogenous p65 nor that of I kappaB alpha and I kappaB beta is altered by low potassium treatment. Similarly, no changes are seen in the nuclear or cytoplasmic levels of p50, p52, RelB and c-Rel. Phosphorylation of p65, which can lead to its activation, is unchanged. Phosphorylation of I kappaB beta is, however, reduced by low potassium treatment. Besides being necessary for high potassium-mediated neuronal survival, NF-kappaB is also involved in the survival-promoting effects of IGF-1 and cAMP as judged by the ability of SN-50 to inhibit the actions of these survival factors and the ability of these factors to inhibit the low potassium-induced alterations in the DNA-binding activity of NF-kappaB. Taken together, our results show that NF-kappaB may represent a point of convergence in the signaling pathways activated by different survival factors and that uncommon mechanisms might be involved in NF-kappaB-mediated survival of cerebellar granule neurons.