The multifaceted role of FOXOs in the maintenance of genomic and metabolic homeostasis

Abstract

FOXO transcription factors are involved in a variety of biological processes including cell cycle regulation, reactive oxygen species (ROS) detoxification and cellular metabolism, ultimately regulating cellular homeostasis. In this thesis we describe novel roles of FOXOs in the maintenance of genomic and metabolic homeostasis. We show that FOXOs are activated in response to DNA double-strand breaks (DSBs) by acquiring DNA damage-induced phosphorylation and then accumulate to the sites of damage. The accumulation of FOXOs to the sites of damage depends on the interaction with components of the MRN complex and it is required for the recruitment of the adaptor protein TRRAP to the DSBs. In this way, FOXOs mediate epigenetic changes around DNA DSBs (histone acetylation), leading to the promotion of DNA-end resection and homologous recombination (HR) repair. HR is an error-free repair mechanism that ensures the maintenance of genomic stability and is thereby an important tumor suppressive mechanism. We also report that this novel function of FOXOs is independent of their role as transcription factors. We further describe a global gene expression analysis performed in cells depleted of FOXO transcription factors. In this analysis, inter alia we identified several metabolic genes whose expression is dependent on FOXOs. One of these genes is isocitrate dehydrogenase 1 (IDH1). IDH1 is a cytosolic enzyme linked to the TCA cycle that mediates the maintenance of α-ketoglutarate and NADPH cytosolic levels. Mutations in IDH1 result in an enzyme with altered enzymatic activity that promotes tumorigenesis via the generation of the oncometabolite 2-hydroxyglutarate and mutant IDH1 is therefore considered an oncogene. We demonstrate that FOXOs are required for the transcription of both the wild-type and the mutant IDH1 and thereby play a dual role in tumorigenesis, by mediating both tumor suppression and tumor promotion. Untransformed cells require FOXO-mediated IDH1 transcription to maintain the levels of NADPH and α-ketoglutarate and thereby regulate ROS scavenging mechanisms and the function of enzymes involved in epigenetic changes. On the other hand, FOXOs are important for the proliferation of cells grown in hypoxia, possibly by promoting IDH1-mediated reductive-carboxylation, and also of transformed cells carrying mutant IDH1, by mediating mutant IDH1 transcription and 2-hydroxyglutarate accumulation. We further show that FOXOs mediate processes linked to mitochondrial health and function. We show that FOXOs are required for the maintenance of cellular bioenergetics by mitochondrial metabolism. We also provide evidence that in the absence of FOXO the expression levels of certain mitochondrial regulators, including TFAM, TDP1, UCP2 and PINK1, are significantly reduced, resulting in the accumulation of mitochondria with elevated mitochondrial DNA damage, increased ROS leakage and decreased turnover. All the above constitute mitochondrial dysfunction, which is linked to aging and aging-associated disorders, for example Parkinson’s disease. In conclusion, our results demonstrate that FOXOs mediate cellular homeostasis in multiple levels. Understanding the upstream regulation of FOXOs that imprints on their functional output will prove particularly beneficial for the treatment of cancer and aging-associated disorders.