Nuclear factor-κb is constitutively active in c-cell carcinoma and required for ret-induced transformation

Specific point mutations of the RET proto-oncogene have been demonstrated to be responsible for multiple endocrine neoplasia (MEN) types 2A and 2B, for familial medullary thyroid carcinoma (MTC) syndromes, as well as for sporadic MTC. Here we show that nuclear factor (NF)- Kappa B is activated in RET-associated C-cell carcinoma specimens. TT cells, a human MTC cell line expressing MEN 2A type RET, display transcriptionally active RelA(p65) in the nucleus. NF- Kappa B activity in these cells is attributable to constitutive I Kappa B kinase (IKK) activity and high turn over of I Kappa B alpha . RET harboring the mutations C634R (MEN 2A) or M918T (MEN 2B), in contrast to wild-type RET, activates a NF- Kappa B-dependent reporter construct upon transient transfection in HeLa cells. We show that the prototpe RET mutation C634R enhances phosphorylation of I Kappa B alpha by IKK beta but not by IKK alpha . RET-induced NF- Kappa B and IKK beta activity requires Ras function but does neither involve the classical mitogen-activated protein kinase kinase/extracellular signal-regulated kinase nor the phosphoinositide 3-kinase/Akt pathways. In contrast, RET-induced NF- Kappa B activity is dependent on Raf and MEKK1. Inhibition of constitutive NF- Kappa B activity results in cell death of TT cells and blocks focus formation induced by oncogenic forms of RET in NIH 3T3 cells. These results suggest that RET-mediated carcinogenesis critically depends on IKK activity and subsequent NF- Kappa B activation.