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On the Regulation and Multiple Functions of the Key Gluconeogenic Enzyme Fbp1 in Rapidly Proliferating Cells: Insights from Yeast and Breast Cancer Cells

Ghanem, Ali

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Abstract

Rewired cancer metabolism sprang into the spotlight this century as a crucial aspect of the malignant transformation. Being the century of the DNA, most 20th century cancer research focused on approaching cancer as a direct result of accumulated mutations while overlooking the metabolic aspects. Albeit very insightful on the origin of the disease, this approach has fallen short of eradicating cancer. Hence efforts in oncology research embarked on exploring other aspects in a bid for a whole-rounded anti-cancer approach. Consequently, last decade has seen accelerating breakthroughs in elucidating the metabolic adaptation of cancer cells to provide the requirements of rapid proliferation and the implication thereof as potential drug targets. Basic metabolic networks and enzymes show large degree of evolutionary conservation between eukaryotes. Moreover, yeast and cancer share the major metabolic hallmark of aerobic glycolysis, demonstrate comparable proliferative tendencies, comparable cell cycle regulation and also undergo apoptosis akin to higher eukaryotes. This multiple overlap makes yeast an attractive model for addressing glucose metabolism as a driver for rapid proliferation. This thesis features the results of my investigation into the regulation and effects of the key gluconeogenic enzyme fructose 1,6 bis-phosphatase Fbp1 in two distinct eukaryotic systems: the budding yeast Saccharomyces cerevisiae and human breast cancer cell lines. Based on previous research pointing out links between Fbp1 and the particular type of DNA damage elicited by methyl methanesulfonate. When exposed to MMS, Fbp1 is transcriptionally upregulated as a part of the response to DNA methylation. Fbp1, on the other hand, bestows an increased MMS-sensitisation upon yeast. I devised a mutational analysis of evolutionary conserved residues to address the mechanism of this additional phenotype, the results of which demonstrated that the enzymatic activity could also be decoupled from further MMS sensitisation, hence suggesting a non-catalytic origin of the additional effect. ii I then embarked on investigating the effects of Fbp1 in breast cancer cell lines of both luminal and basal-like lineages. The findings I present make the case for an anti proliferative role of Fbp1 as an outcome of diminished glucose sensitivity, uptake and a shift from glycolysis to higher mitochondrial activity. Moreover, Fbp1 exhibited a pro oxidative role in cell lines manifested in increased ROS accumulation and sensitivity to oxidative agents. My findings also provide transcriptional evidence of a Wide - scale cell-cycle inhibitory outcome of Fbp1 ectopic expression in Fbp1-deficient MDA-MB231. Intriguingly, I observed that Fbp1-deficient basal like breast cancer cells acquired the capability to post-transcriptionally breakdown ectopic Fbp1 upon long-term stable over-expression. I also demonstrated that this degradation occurs through the proteasome and exclusively upon the long-term selection of cells with ectopic Fbp1. An overview indicates multiple similarities of Fbp1 effects in yeast and cancer cell systems. In general, Fbp1 has comparable effects on proliferation, glycolysis, and Redox balance in both systems. The proteasomal degradation of Fbp1 in cells draws another parallel to the regulation of this enzyme in yeast since the so called “catabolite degradation” of Fbp1 had been long described and well-studied in yeast. Nevertheless, when compared, the two corresponding mutations in yeast and human Fbp1 had distinct consequences for catalytic activity and enzyme stability in each system., therefore hinting at structural differences in the activation mechanisms and at different interaction partners in the two systems. Altogether, the results presented in this thesis endorse Fbp1 as a quasi tumour suppressor and emphasise the potential therapeutic significance of approaches that can de-repress Fbp1 in cancers lacking it and prevent its ablation in cancers that express it.

Item Type: Dissertation
Supervisor: Wölfl, Prof. Dr. Stefan
Date of thesis defense: 25 September 2018
Date Deposited: 18 Oct 2018 08:39
Date: 2018
Faculties / Institutes: The Faculty of Bio Sciences > Institute of Pharmacy and Molecular Biotechnology
Subjects: 000 Generalities, Science
570 Life sciences
Controlled Keywords: Cancer Metabolism, Gluconeogenesis, FBP1
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