14-3-3σ gene silencing during melanoma progression and its role in cell cycle control and cellular senescence

Background: The family of 14-3-3 proteins plays an important role in cancer biology by interfering with intracellular signalling pathways and cell cycle checkpoints. The 14-3-3σ isoform acts as a tumor suppressor and is often inactivated during tumor development. Results: Here, we demonstrate enhanced CpG methylation of the 14-3-3σ gene in lymph node and cutaneous melanoma metastases compared with primary tumors, associated with dramatically reduced mRNA expression. In line with this, treatment of different metastatic melanoma cell lines with 5-aza-2′-deoxycytidine (5-Aza-CdR), a potent inhibitor of cytosine methylation, significantly induces 14-3-3σ protein expression. Additional treatment with histone deacetylase inhibitor 4-phenylbutyric acid (Pba) further enhances 14-3-3σ expression. Induction of 14-3-3σ expression by 5-Aza-CdR/Pba treatment leads to almost complete inhibition of cell proliferation, with cells predominantly arrested in G2-M. The antiproliferative effect of 5-Aza-CdR/Pba was reversed in 14-3-3σ knockdown cells. Similarly, melanoma cell lines stably overexpressing 14-3-3σ show dramatically reduced cell proliferation rates. Moreover, synchronous 14-3-3σ stably overexpressing cells do not progress through cell cycle, but display a permanent increase in the population of 4n DNA containing cells. Interestingly, overexpression of 14-3-3σ induces senescence of melanoma cells and is involved in melanoma cell senescence under genotoxic stress. Finally, 14-3-3σ knockdown supports migratory capacity of melanoma cells in vitro, while 14-3-3σ overexpression has opposing effects. Conclusion: Taken together, the present report indicates that epigenetic silencing of 14-3-3σ might contribute to tumor progression in malignant melanoma via loss of cell cycle control, impaired cellular senescence program and support of migratory capacity.