Abstract
Melanoma is the sixth most common cancer in the USA. Introduction of BRAF inhibitors and advances in cancer immunotherapy has greatly improved treatment of melanoma, however, the problem of tumour relapse and therapy resistance persists. Better understanding of the mechanism of development of the diseases is urgently needed to improve therapeutic intervention. Role of DNA and histone modifications have been well established in several diseases, however, RNA modifications, specifically, N6 methyl adenosine (m6A), has only recently begun to be shown to play critical role in cancer. m6A plays crucial roles in mRNA stability, splicing, export, translation and decay and is regulated by: methyltransferase complex proteins METTL3, METTL14, and WTAP; the demethylases FTO and ALKBH5, and the m6A reader proteins. In the present study, we uncover the role of RNA methyltransferase METTL3 in human melanoma cells. METTL3 expression was assessed in 8 melanoma cell lines (A375, A375MA2, A2058, WM164, 451Lu, WM793, SKMEL2, WM3918). Western blotting data revealed METTL3 to be upregulated in seven melanoma cell lines (except A2058) relative to normal melanocytes. To uncover the role of METTL3 in melanoma, we knockdown METTL3 with lentiviral shRNA and studied the effect on melanoma cells tumorigenesis. The colony formation assay revealed reduced colony number in the METTL3 knockdown A375, A375MA2 and WM793 cells relative to the scrambled control cells. The main cause of melanoma associated fatality is its metastatic dissemination we therefore, next sought to investigate whether METTL3 could affect melanoma cells migration and invasiveness. We performed migration and invasion assays to detect the migratory potential of control or METTL3 knockdown A375, A375MA2 and WM793 cells. Interestingly, both migration and invasion was found to be reduced in METTL3 knockdown cells in all the three cell lines examined. RNA dot blot assay demonstrated a decreased m6A level in total RNA of the METTL3 knockdown cells relative to control cells. p16 and p53 mRNA were found to be enriched in m6A immunoprecipitated samples as compared to IgG control, pointing towards a potential role of m6A in regulating expression of melanoma associated genes. Ongoing effort is aimed at identifying what known or novel regulators of the migratory/invasive cascade is regulated by METTL3 and whether it is through m6A mediated or independent mechanisms. In summary, we have shown that METTL3 is overexpressed in human melanoma and play an important role in metastasis.
Citation Format: Ujwal Dahal, Kang Le, Mamta Gupta. Role of RNA methyltransferase METTL3 in melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3323.
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