1231 UBE2N promotes melanoma growth by maintaining MEK and FRA1 signaling

Abstract

UBE2N is a K63-Ubiquitin conjugase previously characterized as a potential therapeutic target in various immunological diseases and cancer. However, its role in melanoma is unclear. In our present study, we have demonstrated that UBE2N and its essential variant partners, UBE2V1 and UBE2V2 were highly expressed in malignant melanoma. Inhibition of UBE2N and the variants via CRISPR and shRNA-mediated approaches markedly decreased melanoma cell proliferation and subcutaneous tumor growth. By immunostaining, we found that UBE2N loss dramatically reduced the expression of the mesenchymal stem cell markers such as N-cadherin, Nestin and ABCB5. Global phosphopeptide analyses via tandem mass spectrometry identified an array of molecules altered by UBE2N loss. Among these, genes that were upregulated include p53 and p16 tumor suppressors while genes that were downregulated include pMEK, pERK, FRA1, and SOX10 gene regulators previously linked to melanoma growth and invasion. Further, we showed that MEK-inhibition and FRA1 gene silencing inhibited cell proliferation, accompanied by decreased SOX10 expression. Conversely, expression of a constitutively active FRA1 mutant restored the expression of pMEK and SOX10, and rescued anchorage independent growth of cells with UBE2N loss. Lastly, we found that treatment with NSC697923, a small molecular inhibitor of UBE2N, inhibited melanoma cell proliferation in vitro and significantly decreased subcutaneous tumor growth of human melanoma cells in immunodeficient mice. These data demonstrate that UBE2N is essential for melanoma growth, characterizing UBE2N as a novel regulator of the MEK/FRA1/SOX10 signaling cascade and a potential therapeutic target for melanoma.