MECHANISMS OF PROGRAMMED CELL DEATH

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We are presently investigating the mechanisms of programmed cell death PCD by identifying stable structural domains within apoptotic effectors and their targets. Two pathways are currently investigated--the CD95/Fas/APO-1 pathyway and the TNFR-1 pathway. Fas-mediated is dependent on the recruitment of effector molecules to transmit the signal received by the receptor to the PCD enzymes (so-called caspases). The effectors FADD, RIP, Mort1 and RAIDD are all presently under study to identify the stable functional domains utilizing a combination of proteolysis/MALDI mass spectrometry in conjuunction with in vitro and in vivo functional assays of the identified pieces. Subsequent to confirmation of functional domains, the complex structures will be determined using a combination of multi-nuclear NMR and/or X-ray crystallography. In a related pathway, a second receptor system centered around TNFR-1 is being investigated, both for its ability to stimulate death as well as to activate the NF-kappaB pathway. In this regard, the effectors TRADD and the TRAF proteins are under study in an effort to elucidate their opposing effects on the cellular life cycle. Again, the goal is to deconstruct the system into the relevant parts and determine th e structures of the functional complexes.