Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with TRADD

Keratin 8 and 18 (K8/18) are the major components of intermediate filament (IF) proteins of simple or single-layered epithelia. Recent data show that normal and malignant epithelial cells deficient in K8/18 are nearly 100 tim es more sensitive to tumor necrosis factor (TNF)induced cell death. We have now identified human TNF receptor type I (TNFR1)-associated death domain p rotein (TRADD) to be the K18-interacting protein. Among IF proteins tested in two-hybrid systems, TRADD specifically bound K18 and K14, type I acidic) keratins. The COOH-terminal region of TRADD interacted with the coil la of the rod domain of K18. Endogenous TRADD coimmunoprecipitated with K18, and colocalized with K8/18 filaments in human mammary epithelial cells. Overex pression of the NH2 terminus (amino acids 1-270) of K18 containing the TRAD D-binding domain as well as overexpression of K8/18 in SW13 cells, which ar e devoid of keratins, rendered the cel Is more resistant to killing by TNF. We also showed that overexpressed NH2 termini of K18 and K8/18 were associ ated with endogenous TRADD in SW13 cells, resulting in the inhibition of ca spase-8 activation. These results indicate that K18 may sequester TRADD to attenuate interactions between TRADD and activated TNFR1 and moderate TNF-i nduced apoptosis in simple epithelial cells.