Modulation of Cell Fate Decisions To Define Novel Cancer Therapies

<p dir="ltr">Regulation of cell fate is important in health and disease, as when these pathways are disrupted in disease, the mechanisms that govern cell fate are often dysfunctional. p53 is an important transcription factor in determining cell fates and is activated in response to damage. Once a damage threshold is reached the transcriptional activity of p53 determines which cell fate occurs, either senescence or apoptosis. p53 interacts with multiple proteins resulting in reciprocal regulation, notably p53 binds to the Tec family kinase BMX causing reciprocal inhibition, but interaction with the multiple domain pro-apoptotic protein BAK results in BAK activation. Furthermore, BMX negatively regulates BAK via Y108 phosphorylation thus the formation of complexes between BMX, BAK and p53 could regulate cell fate. This thesis aims to understand the interplay between these three proteins and determine the resulting cell fate. I aimed to determine the cellular context behind the formation of the complexes between BMX, BAK and p53, and characterise the binding event between the SH3 domain of BMX and the proline rich domain (PRD) of p53. Furthermore, I determined that BMX did not play a role in the induction of senescence, however, treatment of senescent cells with BMX inhibitor BMX-IN-1 and etoposide in combination selectively reduced cell viability in senescent cells. This suggests that whilst BMX is dispensable for inducing senescence, BMX is important in maintaining the viability of senescent cells. Using cell biology techniques, I showed that p53 and BMX form a complex in untreated and senescent cells that upon apoptosis induction dissociates within an hour. This suggests that the complex formed between BMX and p53 is essential in preventing apoptosis through inhibition of p53’s pro-apoptotic functions. Furthermore, the SH3 domain of BMX binds to both BAK and p53 raising the possibility that BAK and p53 competitively bind to BMX.</p>