Constitutive I kappa B kinase activity correlates with nuclear factor-kappa B activation in human melanoma cells

Constitutive IKK activity associated with increased I kappaB alpha phosphor ylation and degradation contribute to the high level of endogenous nuclear factor-kappaB (NF-kappaB) activation in Hs294T melanoma cells as compared w ith RPE cells (R. L. Shattuck-Brandt and A. Richmond, Cancer Res., 57: 3032 -3039, 1997; M. N. Devalaraja et al., Cancer Res., 59: 1372-1377, 1999). To determine whether this endogenous NF-kappaB activation was characteristic of melanoma, we examined the level of constitutive activation of NF-kappaB in a number of melanoma cell lines. We demonstrate here that eight melanoma cell lines exhibit increased I kappaB kinase (IKK) activity, enhanced phos phorylation of I kappaB alpha and p65, and enhanced nuclear localization of p65/p50 in comparison to normal human epidermal melanocytes. The chemokine s, CXC ligand 1 (CXCL1) and CXCL8, but not CXCL5, are highly expressed in m ost of the melanoma cell lines, suggesting that the constitutive production of chemokines is highly correlated to endogenous NF-kappaB activity. Our f ailure to observe a direct relationship between the fold activation of IKK, CXCL1, or CXCL8 mRNA levels and secretion of these chemokines into the cul ture medium suggest that regulation of chemokine expression also occurs at the posttranscription level of mRNA stability and/or translational control. Moreover, recombinant CXCL1 can directly induce IKK activity in normal hum an epidermal melanocytes in a concentration-dependent manner after up-modul ation of CXCL1 protein expression, whereas inhibition of IKR beta activity results in down-modulation of CXCL1 protein expression. Finally, CXCL1 anti body blocks IKK activity and inhibits the proliferation of melanoma cells t o further support the concept that the constitutive activation of NF-kappaB and autocrine effects of CXCL1 play an important role in the pathogenesis of melanoma.