1. Mutation of the Conserved Threonine 8 within the Human ARF Tumour Suppressor Protein Regulates Autophagy.
- Author
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Fontana, Rosa, Guidone, Daniela, Angrisano, Tiziana, Calabrò, Viola, Pollice, Alessandra, La Mantia, Girolama, and Vivo, Maria
- Subjects
INK4a/ARF locus ,LC3 ,autophagy ,cancer ,cytoskeleton ,Autophagy ,HeLa Cells ,Humans ,Mutation ,Threonine ,Tumor Suppressor Protein p14ARF ,Tumor Suppressor Protein p53 ,Tumor Suppressor Proteins - Abstract
BACKGROUND: The ARF tumour suppressor plays a well-established role as a tumour suppressor, halting cell growth by both p53-dependent and independent pathways in several cellular stress response circuits. However, data collected in recent years challenged the traditional role of this protein as a tumour suppressor. Cancer cells expressing high ARF levels showed that its expression, far from being dispensable, is required to guarantee tumour cell survival. In particular, ARF can promote autophagy, a self-digestion pathway that helps cells cope with stressful growth conditions arising during both physiological and pathological processes. METHODS: We previously showed that ARF is regulated through the activation of the protein kinase C (PKC)-dependent pathway and that an ARF phospho-mimetic mutant on the threonine residue 8, ARF-T8D, sustains cell proliferation in HeLa cells. We now explored the role of ARF phosphorylation in both basal and starvation-induced autophagy by analysing autophagic flux in cells transfected with either WT and ARF phosphorylation mutants by immunoblot and immunofluorescence. RESULTS: Here, we show that endogenous ARF expression in HeLa cells is required for starvation-induced autophagy. Further, we provide evidence that the hyper-expression of ARF-T8D appears to inhibit autophagy in both HeLa and lung cancer cells H1299. This effect is due to the cells inability to elicit autophagosomes formation upon T8D expression. CONCLUSIONS: Our results lead to the hypothesis that ARF phosphorylation could be a mechanism through which the protein promotes or counteracts autophagy. Several observations underline how autophagy could serve a dual role in cancer progression, either protecting healthy cells from damage or aiding cancerous cells to survive. Our results indicate that ARF phosphorylation controls proteins ability to promote or counteract autophagy, providing evidence of the dual role played by ARF in cancer progression.
- Published
- 2022