AbstractEpithelial to mesenchymal transition (EMT) is a biological process in which fully differentiated epithelial cells reversibly dedifferentiate into cells with mesenchymal characteristics. The phenotypic plasticity initiated by EMT is pathological in cancer, where it endows cells with highly aggressive traits that facilitate dissemination, therapeutic resistance, and relapse. EMT also induces the formation of cancer stem cells (CSCs) with tumourigenic and metastatic capabilities that, although only constituting a small proportion of the main tumour bulk, possess the capacity to self-renew and differentiate and enhance therapy resistance. EMT is mediated by a complex network of intracellular signalling cascades that function in a coordinated manner to activate several classes of enzymes that facilitate the progression of EMT downstream. Protein kinase C-θ (PKC-θ) has recently emerged as a key chromatin-tethered mediator of inflammatory signal-mediated breast cancer EMT. PKC-θ regulates the expression of multiple enzymes that are implicated in cancer regulation including members of the dual-specificity phosphatase (DUSP) family of signalling enzymes and the epigenetic enzyme, lysine-specific demethylase 1 (LSD1). DUSPs participate in signal transduction cascades where they catalyse the dephosphorylation of threonine/serine and tyrosine residues on their target substrates. LSD1 is a key histone demethylase that alters the epigenetic landscape by selectively catalysing the demethylation of mono- and di-methylated H3K4 and H3K9, subsequently facilitating gene repression and activation, respectively. This thesis examined the contribution of several members of the DUSP family, namely DUSP1, DUSP4 and DUSP6, as well as LSD1 in the regulation of breast cancer EMT and the plasticity of CSCs.
Work in this thesis shows DUSP1, DUSP4, DUSP6 and LSD1 are induced in response to PKC pathway-mediated activation of EMT. DUSP1 and DUSP4 globally exist with distinct histone post-translational modifications and directly tether to the chromatin-template at mesenchymal gene loci. Specifically, DUSP4 globally regulates the acetylation of H3K27 by modulating the activity of the key histone acetyltransferase, p300. Furthermore, DUSP1, DUSP4 and DUSP6 regulate the plasticity of breast CSCs where they play differential roles in their formation. Induction of LSD1 promotes the progression of EMT where it subsequently regulates the expression of the key EMT markers, E-cadherin, vimentin and Snail. Chromatin anchored PKC-θ phosphorylates LSD1 at serine-111 (LSD1-s111p). This phosphorylation event is critical for LSD1’s demethylase and EMT promoting activity and LSD1-s111p is enriched in chemoresistant cells in vivo. In turn, LSD1 induces pan-genomic gene expression in networks implicated in EMT and CSC regulation. At the genome-wide level, LSD1 selectively induces gene expression programs in CSCs whilst repressing non-CSCs programs. Furthermore, LSD1 couples to PKC-θ on the mesenchymal gene epigenetic template where it promotes LSD1-mediated gene induction. Collectively, these results provide novel insight into the multi-layered regulation of breast cancer EMT and characterises enzymes that would potentially serve as promising epigenetic targets for adjuvant anti-cancer therapy.
|Date of Award||2019|
|Supervisor||Sudha Rao (Supervisor) & Anjum Zafar (Supervisor)|