TRADD-dependent signaling and the interaction of TRADD with death domain-containing proteins

Members of the TNF receptor superfamily share a homologous region termed a death domain that mediates recruitment of an adaptor protein to the receptor facilitating the activation of downstream signaling. The adaptor proteins recruited to the receptors also possess a death domain and this region is required for binding to the receptor. The primary adaptor protein for TNFR1 signaling is TRADD. TRADD is comprised of two distinct domains, a death domain and a TRAF-binding domain. While the TRAF domain mediates recruitment of TRAF-2 to promote JNK activation, the death domain is required for the propagation of apoptotic signaling. Our lab showed that TRADD contains both a bipartite nuclear localization sequence that lies within the death domain, as well as a CRM-1 dependent nuclear export sequence. Further we found that accumulation of TRADD in the nucleus by inhibition of export or by expression of the truncated TRADD death domain, activated an apoptotic pathway dependent upon nuclear localization of TRADD. In characterizing the apoptotic pathway activated by nuclear TRADD we determined that death was dependent upon caspase as well as serine protease activation. In addition, we show that caspase 9 is required for the activation of the death pathway, but that caspase 9 can be activated in an APAF-1 independent mechanism.; Apoptosis activated in response to both Fas Ligand as well as TNF-alpha requires the adaptor protein FADD. Like TRADD, FADD is composed of two domains, a death domain that mediates binding to both the Fas receptor as well as to TRADD, and a death effector domain that recruits caspase 8 to the DISC. Significant mutational analysis has been performed on the FADD-death domain, which is comprised of six alpha helices, and these studies indicate that helices 2 and 3 are important in binding to Fas and TRADD. Recently new data emerged indicating that an expanded binding surface on the FADD death domain facilitated FADD/Fas interactions. Using a series of point mutations in the FADD death domain we tested whether this expanded surface also played a role in FADD/TRADD interaction. We found that there are specific amino acid residues within FADD's death domain that are required for interaction with TRADD compared with Fas and vice versa. Based upon our work and the work of others, it can be concluded that death domain interactions are more complex than the presence of the homologous sequence and that different death domains require different requirements to facilitate various interactions. (Abstract shortened by UMI.)...