A novel NF-κB-inducing kinase-MAPK signaling pathway up-regulates NF-κB activity in melanoma cells

Constitutive activation of NF-κB is an emerging hallmark of various types of tumors including breast, colon, pancreatic, ovarian, and melanoma. In melanoma cells, the basal expression of the CXC chemokine, CXCL1, is constitutively up-regulated. This up-regulation can be attributed in part to constitutive activation of NF-κB. Previous studies have shown an elevated basal IκB kinase (IKK) activity in Hs294T melanoma cells, which leads to an increased rate of IκB phosphorylation and degradation. This increase in IκB-α phosphorylation and degradation leads to an ∼19-fold higher nuclear localization of NF-κB. However, the upstream IKK kinase activity is up-regulated by only about 2-fold and cannot account for the observed increase in NF-κB activity. We now demonstrate that NF-κB-inducing kinase (NIK) is highly expressed in melanoma cells, and IKK-associated NIK activity is enhanced in these cells compared with the normal cells. Kinase-dead NIK blocked constitutive NF-κB or CXCL1 promoter activity in Hs294T melanoma cells, but not in control normal human epidermal melanocytes. Transient overexpression of wild type NIK results in increased phosphorylation of extracellular signal-regulated kinases 1 and 2 (ERK1/2), which is inhibited in a concentration-dependent manner by PD98059, an inhibitor of p42/44 MAPK. Moreover, the NF-κB promoter activity decreased with overexpression of dominant negative ERK expression constructs, and EMSA analyses further support the hypothesis that ERK acts upstream of NF-κB and regulates the NF-κB DNA binding activity. Taken together, our data implicate involvement of IκB kinase and MAPK signaling cascades in NIK-induced constitutive activation of NF-κB.