PO-254 Merkel cell polyomavirus T-antigen regulate microRNAs post-transcriptionally through DICER in merkel cell carcinoma

Abstract

IntroductionBackground and aimMerkel cell carcinoma (MCC) is an aggressive type of skin cancer. About 80% of MCCs harbour integrated Merkel cell polyomavirus (MCV) genome with a mutation in the large T (LT) gene. MCV T-antigens are required for neoplastic transformation and maintenance of cell growth. however the molecular mechanism by which the virus induces tumorigenesis remains unclear. In this study we aimed to identify and characterise functional role of MCV-regulated microRNAs in MCC.Material and methodsWe did silencing or ectopic expression of MCV T-antigens to identify specific miRNAs regulated by MCV T-antigens by RT-qPCR. The involvement of MCV T-antigens and miRNAs in autophagy was evaluated using LC3-II conversion, mRFP-EGFP-LC3 reporter and/or transmission electron microscopy. The targets of miRNAs were verified by western blot analysis and luciferase reporter assays. We also look for the DICER expression by western blot.Results and discussionsWe identified specific miRNAs regulated by MCV T-antigens. Using both gain- and loss-of-function experiments, we showed that MCV T-antigens and MCV-regulated miRNAs (miR-375, miR-30a-5p and miR-30a-3p) could regulate autophagy. We further demonstrated that miR-375 could directly regulate ATG7 and SQSTM1, while both miR-30a-3p and miR-30a-5p could target BECN1. We also identified the MCV T-antigens regulates these miRNAs through post-transcriptionally by regulating DICER expression in MCV positive MCC.ConclusionWe provide evidence that MCV T-antigens regulate miRNAs and autophagy in MCC. MCV T-antigens increase miRNA expressions through pots-transcriptional regulation, and several of these miRNAs can directly target multiple autophagy genes that lead to suppression of autophagy in MCC.