Pharmacological activation of SIRT6 triggers lethal autophagy in human cancer cells

Sara Iachettini,Clemens Steegborn,Antonello Mai,Erica Salvati,Angela Rizzo,Annamaria Biroccio,Alessia Lucidi,Pasquale Zizza,Dante Rotili,Donatella Del Bufalo,Daniela Trisciuoglio,Luca Di Leo,Carlo Leonetti,Maria Rosa Ciriolo

doi:10.1038/s41419-018-1065-0

Abstract

Sirtuin 6 (SIRT6) is a member of the NAD+-dependent class III deacetylase sirtuin family, which plays a key role in cancer by controlling transcription, genome stability, telomere integrity, DNA repair, and autophagy. Here we analyzed the molecular and biological effects of UBCS039, the first synthetic SIRT6 activator. Our data demonstrated that UBCS039 induced a time-dependent activation of autophagy in several human tumor cell lines, as evaluated by increased content of the lipidated form of LC3B by western blot and of autophagosomal puncta by microscopy analysis of GFP-LC3. UBCS039-mediated activation of autophagy was strictly dependent on SIRT6 deacetylating activity since the catalytic mutant H133Y failed to activate autophagy. At the molecular level, SIRT6-mediated autophagy was triggered by an increase of ROS levels, which, in turn, resulted in the activation of the AMPK-ULK1-mTOR signaling pathway. Interestingly, antioxidants were able to completely counteract UBCS039-induced autophagy, suggesting that ROS burst had a key role in upstream events leading to autophagy commitment. Finally, sustained activation of SIRT6 resulted in autophagy-related cell death, a process that was markedly attenuated using either a pan caspases inhibitor (zVAD-fmk) or an autophagy inhibitor (CQ). Overall, our results identified UBCS039 as an efficient SIRT6 activator, thereby providing a proof of principle that modulation of the enzyme can influence therapeutic strategy by enhancing autophagy-dependent cell death.

Highlights

Sirtuins are histone deacetylase enzymes that use nicotinamide adenine dinucleotide (NAD+) as a cosubstrate for their enzymatic activities
In particular deacetylation of histone H3K9 was found in HeLa epithelial cervix carcinoma and HCT116 colon carcinoma lines, while no effect was observed in HT1080 human fibrosarcoma cell line expressing a higher basal level of H3K9 histone variant (Supplementary 1A)
The role of Sirtuin 6 (SIRT6) on autophagy-mediated cell death was corroborated by a genetic approach demonstrating that overexpression of wild-type, but not the catalytic inactive form of SIRT6 was able to increase cell death that was reverted by CQ treatment (Fig. 6c). These results clearly demonstrate that pharmacological activation of SIRT6 triggers an increase of autophagy that results in apoptosis in different human cancer cells

Summary

Introduction

Sirtuins are histone deacetylase enzymes that use nicotinamide adenine dinucleotide (NAD+) as a cosubstrate for their enzymatic activities. They are mainly involved in regulation of cell stress response and metabolism, playing key roles in normal and cancer cells[1]. Among the components of mammalian sirtuin family, SIRT6 deacetylates the histone H3 on acetylated K9, K562,3, and the more recently identified K18 and K27 residues[4,5], causing the repression of many genes involved in inflammation, aging, genome stability, metabolic pathways, and telomere integrity[2,6,7,8].

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