SIRT1 is an NAD-dependent deacetylase which influences the outcome of cellular stress by regulating histone-mediated chromatin remodelling and the p53-dependent DNA damage response. As such, SIRT1 modulates key signalling pathways which influence carcinogenesis. SIRT1 expression is regulated by p53 activity at the transcriptional level, and by the microRNA miR-34a at the translational level. Together SIRT1, p53 and miR-34a form a feedback loop which self-regulates p53 activation.
The tumour suppressor p53 is mutated in >60% of all cancers, including those affecting the skin. Thus the development of pharmaceutical agents which differentially target cells on the basis of p53 mutational status is of interest in the prevention and treatment of skin cancer. In this study, we used primary keratinocytes (neonatal human epidermal keratinocytes - NHEKs) and p53-mutated keratinocytes (HaCaTs) cultured in identical serum-free conditions to investigate the relationship between p53 mutational status, SIRT1 activity and miR-34a processing in human keratinocytes.
Expression of miR-34a was up-regulated by transient ablation of SIRT1 in NHEKs; but was suppressed by dual knockdown of SIRT1 and p53. In contrast, SIRT1 knockdown reduced miR-34a in HaCaTs keratinocytes whilst concomitantly up-regulating expression of its precursor transcript. This discrepancy between mature and precursor transcript levels was also observed after pharmaceutical SIRT1 inhibition in both p53-ablated and HaCaT keratinocytes. Additionally, differences in the level of p53 expression and acetylation observed between the NHEK and HaCaT cell lines was partially restored by inhibiting SIRT1 in the p53-mutated cell line.
Our results indicate that p53 haploinsufficiency impairs regulatory control of the SIRT1-miR-34a axis. SIRT1 inhibitors enable keratinocytes with p53 mutations to regulate p53 expression and relative acetylation and may therefore have therapeutic potential for the treatment and prevention of skin cancer.