The Identification and Characterisation of Novel Proteins and Metabolic Signalling Pathways that Regulate TRAIL-Induced Apoptosis

TNF-related apoptosis inducing ligand (TRAIL) is a putative anti-cancer cytokine that is relatively specific for transformed cells however Mantle Cell Lymphoma (MCL) is resistant. TRAIL induces apoptosis through formation of a Death-Inducing Signalling Complex (DISC) and the successive activation of a caspase cascade. Therefore, the aims of this thesis were two-fold: firstly to examine the known and novel protein components of the DISC through mass spectrometry analysis and second to determine how cellular metabolism, specifically glucose-deprivation, influences TRAIL-induced apoptosis. A tagged variant of TRAIL was used to capture the DISC from a TRAIL-sensitive MCL-derived cell line (Z138) allowing for its purification and subsequent proteomic analysis by mass spectrometry. From this study, it was found that the transferrin receptor (TFR1) may interact with the DISC, specifically with the TRAIL-receptors, where it potentially modulates TRAIL-induced apoptosis and/or receptor-mediated endocytosis. In addition, it was observed that the known DISC component FADD was present in sub-stoichiometric amounts compared to caspase-8 and this permitted the identification of a novel DISC structure based on caspase-8 chain formation. In parallel to this, glucose-deprivation studies were performed, which showed that, in the absence of glucose, Z138 cells was less sensitive to TRAIL-induced apoptosis. Furthermore, these cells also showed a reduced sensitivity to both ABT-737 and x-ray radiation. As a result, proteomic, ultra-structural and gene profiling studies were performed on Z138 cells cultured in the absence of glucose, which demonstrated a switch towards an anti-apoptotic/pro-survival phenotype.