J Biol Chem. 1996 Apr 19;271(16):9858-62.

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

Park, A., Baichwal, V. R.,
--- - Tularik Inc., South San Francisco, California 94080, USA.
Tumor necrosis factor receptor 1 (TNF-R1) mediates most of the biological properties of TNF including activation of the transcription factor NF-kappaB and programmed cell death. An approximately 80-amino acid region within the intracellular domain of the receptor, termed the death domain, is required for signaling NF-kappaB 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-kappaB 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-kappaB 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-kappaB was found to act as dominant negative mutant capable of preventing induction of NF-kappaB by TNFalpha. Such observations provide evidence that TRADD performs an obligate role in TNF-induced NF-kappaB activation.
PMID: 8621670External
Arrow2 In vitro interaction Arrow2 In vivo interaction Arrow2 Characterization Arrow2 Functional role Arrow2 top
In vitro interaction
  DD1 DD2 Reference
Family DD1 DD2 Method Species Region Expression Species Region Expression
DD TNFR1 Link TRADD GST fusion protein pull-down Human 214-426 Not specified Human 196-312 In vitro translation 8621670
DD TRADD Link TRADD GST fusion protein pull-down Human 196-312 In vitro translation Human 214-426 Not specified 8621670
(Link: click this icon to show interactions only between the two corresponding DDs)
Arrow2 In vitro interaction Arrow2 In vivo interaction Arrow2 Characterization Arrow2 Functional role Arrow2 top
In vivo interaction
  Endogenous
expression
Overexpression DD1 DD2 Reference
Family DD1 DD2 Method Species Region Species Region
DD TNFR1 Link TRADD Co-immunoprecipitation HEK293 Not specified Not specified Human 112-312 8621670
DD TRADD Link TRADD Co-immunoprecipitation HEK293 Human 112-312 Human 1-312 8621670
(Link: click this icon to show interactions only between the two corresponding DDs)
Arrow2 In vitro interaction Arrow2 In vivo interaction Arrow2 Characterization Arrow2 Functional role Arrow2 top
Characterization
  Stoichiometry Affinity Protein-Protein interface Reference
Family DD1 DD2 Method Mutation Complex structure
DD TNFR1 Link TRADD GST fusion protein pull-down 196-312 substituted A(TRADD) 8621670
(Link: click this icon to show interactions only between the two corresponding DDs)