Programmed cell death (PCD) is a carefully controlled process that is vital for the proper development of an organism as well as for maintaining its homeostasis. However, when the process is misregulated autoimmune disorders, neurodegenerative diseases, and cancer can develop. Understanding the molecular mechanisms of PCD is crucial to understanding how these diseases develop and will likely provide insights into therapeutic drugs. Key to the process of PCD is the activation of the cell death proteases called caspases; caspase activation rapidly causes cell disassembly. However, the in vivo molecular targets of caspases are largely unknown; it is likely that these targets are responsible for distinct cell disassembly events. C. elegans provides the ideal system in which to identify the downstream targets of its caspase, CED-3. Using a sensitized CED-3 GFP suppressor screen two putative downstream CED-3 targets have been identified: cps-1 and -3.
The specific aims of this proposal are 1) to genetically (e.g. three factor mapping, complementation tests, and epistasis analysis) and phenotypically (cell corpse counts, cell structure specific GFP markers) characterize these two mutants in order to establish if they are bona fide cell death mutants; 2) to clone cps-1 and -3 using cosmid rescue; 3) to biochemically and functionally analyze cps-1 and -3 using a protease assay in order to determine if they are cleavage targets of CED-3; and finally, the long term goal, 4) to conduct a cps-3 enhancer screen to identify additional molecules involved in cell disassembly.
