| The underlying mechanism of cell death involves a complex network of biological components which orchestrate specific biochemical events. It is a well-conserved process, revealed by sophisticated genetic tools available in model organisms such as C. elegans, Drosophila and mice. Furthermore, these model organisms have also helped discover different alternative mechanisms, particularly ones that deviate from the well-characterized caspase-dependent pathway. This dissertation aims to understand two outstanding questions in the area of cell death: (1) Is the mitochondrial endonuclease G (EndoG) a bonafide death effector? (2) Does PARP-1 cleavage by caspases play a role in cell death? To address these questions, we generated two mutant mouse strains: (1) a knockout (KO) of EndoG; and (2) a mutant PARP-1 (PARP-1 KI) that is resistant to caspase cleavage. Interestingly, both EndoG KO and PARP-1 KI mice were healthy and viable. To study the role of EndoG and cleavage of PARP-1 in various cell death paradigms, we established mouse embryonic fibroblasts (MEFs) from these mice and subjected them to various toxic stimuli that represent different cell death pathways. We found that both EndoG KO and PARP-1 KI MEFs were equally susceptible to death as wild-type (WT) MEFs. These mutant mice were also equally sensitive to excitotoxic stress as WT mice. Our results suggest that EndoG and PARP-1 cleavage by caspases are dispensable for cell death. However, given the robustness of the cell death pathway with its molecular players having redundant or overlapping functions, perturbing a single component may not be enough to disrupt cell death. These mouse models, therefore, are valuable tools for future studies perturbing several related events, rather than singly, to be able to uncover mechanisms of cell death. |
