ROLE OF POST-TRANSLATIONAL MODIFICATIONS IN MODULATING APE1 FUNCTIONS IN TUMOUR CELLS

Apurinic/apyrimidinic endonuclease 1 (APE1) is the main mammalian endonuclease involved in the repair of DNA lesions caused predominantly by oxidative and alkylating stresses through its participation in base excision repair (BER) pathway. Although APE1 was discovered for its ability to cleave and remove AP-sites and to enhance the DNA binding properties of several cancer-related transcription factors, through redox-dependent mechanisms (involving its so-called Ref-1 activity), in the latest years investigators have described new and broader functions for this DNA repair enzyme. In fact, it has been demonstrated a direct role for APE1 in the regulation of gene transcription and an unexpected role in the RNA metabolism being able to cleave damaged or site-specific RNAs. Despite different works regarding the transcriptional and post-translational mechanisms that cells used to control and to redirect APE1 to its several functions, it is still a matter of debate the role of the first 33 amino acids, a unique evolutionary N-terminus specific for the mammalian protein, that impacts, through macromolecules interaction and post-translational modifications, on controlling APE1 activities. In this work of Thesis, new acetylated lysine residues were identified in vivo and the role of acetylation sites at N-terminus (Lys 27-35) in regulating APE1 functions and subcellular localization were studied. Moreover, seventeen new interacting partners were identified. Among these, the attention was focused on APE1 and Nucleophosmin 1 (NPM1) interaction within nucleoli and nucleoplasm. NPM1 is a nucleolar protein, mainly involved in ribosome biogenesis, stress responses and genome maintenance. Growing body of evidences emphasizes a role for NPM1 in DNA repair field, but its exact role(s) has not been identified yet. Interestedly, patients with acute myeloid leukemias (AMLs), characterized by the expression of a mutated form of NPM1 (NPM1c+), causative for its aberrant cytoplasmic localization, represent better responders to chemotherapy and for favorable overall survival. At present, the molecular reasons underneath the role for NPM1 in tumorigenesis of solid tumors and in AMLs are still lacking. In this framework, a clear contribution of NPM1 in DNA repair control, through the functional regulation of the APE1 endonuclease activity in BER pathway, its subcellular localization and stability has been demonstrated in vivo. In this light, the positive clinical impact of NPM1c+ in chemotherapeutic response might be related to the potential interference with the functions of NPM1 interacting partners, once delocalized in the cytoplasm. An intriguing mechanism for explaining also the biological effects of APE1 genetic variants, considered in this work of Thesis, supported by the observation that the majority of the polymorphisms presents an altered complex network of interactions, protein stability and stress response, affecting the APE1 functional status. Our findings provide a glimpse into the role of the nucleolus and NPM1 in controlling APE1 functions, suggesting a critical role for the intricate network of APE1 interacting partners, especially NPM1, and post-translational modifications in BER in vivo that might be important for explaining the APE1 dysregulation seen in different types of tumors.