Targeted deletion of MKK4 gene potentiates TNF-induced apoptosis through the down-regulation of NF-κB activation and NF-κB-regulated antiapoptotic gene products

MAPK kinase 4 (MKK4) is a dual-specificity kinase that activates both JNK and p38 MAPK. However, the mechanism by which MKK4 regulates TNF-induced apoptosis is not fully understood. Therefore, we used fibroblasts derived from MKK4 gene-deleted (MKK4-KO) mice to determine the role of this kinase in TNF signaling. We found that when compared with the wild-type cells, deletion of MKK4 gene enhanced TNF-induced apoptosis, and this correlated with down-regulation of TNF-induced cell-proliferative (COX-2 and cyclin D1) and antiapoptotic (survivin, IAP1, XIAP, Bcl-2, Bcl-x_L, and cFLIP) gene products, all regulated by NF-κB. Indeed we found that TNF-induced NF-κB activation was abrogated in MKK4 gene-deleted cells, as determined by DNA binding. Further investigation revealed that TNF-induced IκBα kinase activation, IκBα phosphorylation, IκBα degradation, and p65 nuclear translocation were all suppressed in MKK4-KO cells. NF-κB reporter assay revealed that NF-κB activation induced by TNF, TNFR1, TRADD, TRAF2, NIK, and IκBα kinase was modulated in gene-deleted cells. Overall, our results indicate that MKK4 plays a central role in TNF-induced apoptosis through the regulation of NF-κB-regulated gene products.