Novel functions of the p14ARF tumor suppressor: deciphering cell fate at the crossroad of adhesion and survival

The p14ARF tumor suppressor is an important sensor of oncogenic stimuli inducing cell cycle arrest and or apoptosis through both p53-dependent and independent pathways. It is widely demonstrated that ARF prevalent nucleo/nucleolar localization plays an important role in its tumor suppressor functions. Nevertheless, recent studies show that, despite its role in growth suppression, ARF is overexpressed in a significant fraction of human tumors, suggesting that it may possess a pro-survival function. Accordingly, in our lab it has been demonstrated that, following PKC stimulation, ARF protein is phosphorylated and accumulates in the cytoplasm where it is unable to efficiently control cell proliferation both in tumor and immortalized cell lines. These observations led to the hypothesis that ARF expression in cancer cells could aid tumor progression by conferring pro-survival properties to the cells. On the other hands, ARF cytoplasmic localization could highlight physiologic and novel roles of this versatile protein. In this scenario, the aim of my studies was to better understand ARF functions in HeLa, HaCaT and H1299 cell lines, where ARF is highly expressed and mainly localizes in the cytoplasm. We unexpectedly found that ARF plays a role in cytoskeleton organization. Interestingly, during cytoskeleton remodelling induced by cell spreading, we observed a strong PKC activation resulting in an ARF protein levels increase and stabilization in the cytoplasm. Moreover, we demonstrated that, by aiding cytoskeleton assembly during spreading, ARF protects cells from anoikis blocking DAPK (Death Associated Protein Kinase) dependent apoptosis and promoting cell survival through FAK stabilization. Interestingly, we found that the ARF T8D mutant, mimicking the phosphorylation status of the protein, is able to mediate both cell spreading, FAK activation and cell proliferation through inhibition of DAPK functions. On this basis, we propose that the role of ARF in cell adhesion and survival strictly depends on both cell context and post-translational modifications.