miRNomes of haematopoietic stem cells and dendritic cells identify miR-30b as a regulator of Notch1

Su, Xiaoping; Qian, Cheng; Zhang, Qian; Hou, Jin; Gu, Yan; Han, Yanmei; Chen, Yongjian; Jiang, Minghong; Cao, Xuetao

Stage-specific miRNAs undergo epigenetic reprogramming. Although many miRNA promoters had been identified by using either a computational or experimental approach, numerous miRNA promoters still remain unclear. This difficulty persecuted the researches of epigenetic regulation for miRNAs. The regions (2,500 bp upstream, 500 bp downstream of pre-miRNA) were postulated as promoters of miRNAs to investigate the epigenetic regulation of miRNA expression 25 potential promoter to investigate the epigenetic regulation of stage-specific miRNAs during DC development and differentiation. Acetylation of histone was a general marker associated with actively transcribed promoter 26 , and acetylation of histone in the promoter regions of stage-specific miRNAs were assayed by chromatin immunoprecipitation (ChIP) assay. For miRNA expression, miR-1 and miR-486 were highly expressed in HSCs, whereas poorly expressed in other three stages; miR-132 and miR-147 were highly expressed in imDCs and maDCs, whereas poorly expressed in HSCs and DCreg; and miR-125a-5p and miR-99a were highly expressed in DCreg, whereas poorly expressed in other three stages (Fig. 3a) . These results were confirmed by qRT-PCR assays (Fig. 3b) , and pri-miRNA transcripts of these miRNAs were also confirmed by qRT-PCR (Fig. 3c ). ChIP assays on the potential promoters of the above stage-specific miRNAs were analysed using two sets of qPCR primers spanning the promoters (Fig. 3d) , and results showed that acetylation of histone H3 in the promoter regions of miR-1-1 and miR-486 were high only in HSCs; acetylation of histone H3 in the promoter regions of miR-132 and miR-147 were high only in imDCs and maDCs, whereas acetylation of histone H3 in the promoter regions of miR-99a and miR-125a were high only in DCreg (Fig. 3e) , which were consistent with the stage-specific expression of these miRNAs. Our results suggest that stagespecific miRNAs undergo epigenetic reprogramming during cell development and differentiation.