Targeting the coding sequence: opposing roles in regulating classical and non-classical MHC class I molecules by miR-16 and miR-744

• Published in: Journal for immunotherapy of cancer 2020-06, Vol.8 (1), p.e000396
• Main Authors: Friedrich, Michael, Vaxevanis, Christoforos K, Biehl, Katharina, Mueller, Anja, Seliger, Barbara
• Format: Article
• Language: English
• Subjects: Antibodies; Antigens; Apoptosis; Basic Tumor Immunology; Cancer; Cell Proliferation; Cytotoxicity; Flow cytometry; Gene expression; Gene Expression Regulation, Neoplastic; Histocompatibility Antigens Class I - classification; Histocompatibility Antigens Class I - genetics; Histocompatibility Antigens Class I - immunology; HLA; Humans; Immunotherapy; Laboratories; MicroRNAs; MicroRNAs - genetics; molecular biology; Neoplasms - genetics; Neoplasms - immunology; Neoplasms - pathology; oncology; Proteins; RNA, Messenger - genetics; RNA, Messenger - immunology; Tumor Cells, Cultured
• ISSN: 2051-1426; 2051-1426
• DOI: 10.1136/jitc-2019-000396

BackgroundTo control gene expression, microRNAs (miRNAs) are of key importance and their deregulation is associated with the development and progression of various cancer types. In this context, a discordant messenger RNA/protein expression pointing to extensive post-transcriptional regulation of major histocompatibility complex (MHC) class I molecules was already shown. However, only a very limited number of miRNAs targeting these molecules have yet been identified. Despite an increasing evidence of coding sequence (CDS)-located miRNA binding sites, there exists so far, no detailed study of the interaction of miRNAs with the CDS of MHC class I molecules.MethodsUsing an MS2-tethering approach in combination with small RNA sequencing, a number of putative miRNAs binding to the CDS of human leukocyte antigen (HLA)-G were identified. These candidate miRNAs were extensively screened for their effects in the HLA-G-positive JEG3 cell line. Due to the high sequence similarity between HLA-G and classical MHC class I molecules, the impact of HLA-G candidate miRNAs on HLA class I surface expression was also analyzed. The Cancer Genome Atlas data were used to correlate candidate miRNAs and HLA class I gene expression.ResultsTransfection of candidate miRNAs revealed that miR-744 significantly downregulates HLA-G protein levels. In contrast, overexpression of the candidate miRNAs miR-15, miR-16, and miR-424 sharing the same seed sequence resulted in an unexpected upregulation of HLA-G. Comparable results were obtained for classical MHC class I members after transfection of miRNA mimics into HEK293T cells. Analyses of The Cancer Genome Atlas data sets for miRNA and MHC class I expression further validated the results.ConclusionsOur data expand the knowledge about MHC class I regulation and showed for the first time an miRNA-dependent control of MHC class I antigens mediated by the CDS. CDS-located miRNA binding sites could improve the general use of miRNA-based therapeutic approaches as these sites are highly independent of structural variations (e.g. mutations) in the gene body. Surprisingly, miR-16 family members promoted MHC class I expression potentially in a gene activation-like mechanism.