Systematic mutational analysis of the death domain of the tumor necrosis factor receptor 1-associated protein TRADD

Tumor necrosis factor receptor 1 (TNF-R1) mediates most of the biological properties of TNF including activation of the transcription factor NF-kappa B and programmed cell death, An similar to 80 amino acid region within the intracellular domain of the receptor, termed the death domain, is required for signaling NF-kappa B activation and cytotoxicity, A TNF-R1-associated protein TRADD has been discovered that interacts with the death domain of the receptor, Elevated expression of TRADD in cells triggers both NF-kappa B activation and programmed cell death pathways, The biological activities of TRADD have been mapped to a 111-amino acid region within the carboxyl-terminal half of the protein, This region shows sequence similarity to the death domain of TNF-R1 and can self-associate and bind to the TNF-R1 death domain, We have performed an alanine scanning mutagenesis of TRADD's death domain to explore the relationship among its various functional properties, Mutations affecting the different activities of TRADD do not map to discrete regions but rather are spread over the entire death domain, suggesting that the death domain is a multifunctional unit, A mutant that separates cell killing from NF-kappa B activation by the TRADD death domain has been identified indicating that these two signaling pathways diverge with TRADD. Additionally, one of the TRADD mutants that fails to activate NF-KB was found to act as dominant negative mutant capable of preventing induction of NF-kappa B by TNF alpha. Such observations provide evidence that TRADD performs an obligate role in TNF-induced NF-kappa B activation.