microRNA expression changes in DICER-KO B cells after OVA-specific immune synapse formation

Intercellular communication orchestrates effective immune responses against disease-causing agents. Extracellular vesicles (EVs) are potent mediators of cell-cell communication. EVs carry bioactive molecules, including microRNAs, which modulate gene expression and function in the recipient cell. Here, we show that formation of cognate primary OVA-specific T-B lymphocyte immune contacts promotes transfer to the B cell of a very restricted set of OTII-derived CD4+ T cell exosomal microRNAs (mmu-miR20a-5p, mmu-miR-25-3p and mmu-miR-155-3p). To discriminate exogenously transferred microRNAs from microRNAs induced in B cells after immune synaps, small RNAseq of CD19Cre/Ki DICERfl/fl (DICER-KO) B cells, virtually lacking mature microRNAs, alone and cocultured with OTII CD4+T cells in the presence and absence of OVA was performed; (N=3) independent experiments B cells isolated from CD19Cre/Ki DICERfl/fl (DICER-KO) mice were either left alone or cocultured with OVA-specific OTII-derived CD4+T cells in a 1B:4T ratio, either in the presence or absence of OVA peptide. After 16h culture, B lymphocytes were sorted and total RNA was isolated using the miRNeasy RNA Isolation Kit (QIAGEN). RNA was amplified and prepared for small RNA-Seq using the SmallRNA NEBNext Library Prep Set (New England Biolabs) following the manufacturer’s recommendations and sequenced using the Illumina HiSeq 2500 System. Three biological replicates were analyzed.