Επίδραση μερικής και ολικής καταστολής του διμερούς ARF-μονοπατιού απόκρισης στις δικλωνικές θραύσεις του DNA στην εξέλιξη του καρκίνου: σημασία στη θεραπευτική προσέγγιση

Abstract

The DNA damage response (DDR) pathway and ARF function act as barriers of human cancer development. It has been considered that the DDR and ARF exert this function independently of each other. However, a few studies propose that ARF’s activity is positively regulated by the DDR pathway. Examining this hypothesis we performed a series of experiments using molecular techniques such as immunoblotting, immunohistochemistry, immunofluorescence, immunoprecipitation, Real Time Reverse Transcritpion polymerase chain reaction (RT-PCR) in biological material from cell culture or histological samples, as well as ectopic protein expression through plasmid transfections, proteomic analyses, ribosome RNA biogenesis assay and xenografts of human cancer cells. We surprisingly found that ATM suppressed, in a transcription-independent manner, ARF protein levels and activity. Specifically, ATM activated protein phosphatase 1 (PP1). PP1 antagonized Nek2-dependent phosphorylation of nucleophosmin (NPM), liberating ARF from NPM and rendering it susceptible to degradation by the ULF E3-ubiquitin ligase. In human clinical samples, loss of ATM expression correlated with increased ARF levels and in xenograft and tissue culture models, inhibition of ATM stimulated the tumour-suppressive effects of ARF. The importance of the proposed mechanism can be exploited through a therapeutic approach, especially in cases of tumours bearing loss of p53. In such tumours, DDR may act in favour of the tumour cells, since the major effector of the antiumour barriers of apoptosis and senescence is absent, but ATM inhbition could boost ARF’s tumour-suppressive function, contributing to an anti-tumour response.