Identification of nuclear-enriched miRNAs during mouse granulopoiesis

Abstract

BackgroundMicroRNAs (miRNAs) are coordinators of cellular differentiation, including granulopoiesis. Although differential expression of many miRNAs is associated with the maturation of granulocytes, analysis of differentially expressed miRNAs and their cellular localization across all stages of granulopoiesis, starting from hemopoietic stems cells, is not well characterized.MethodsWe analyzed whole cell miRNA and mRNA expression during granulopoiesis using Taqman low-density and Affymetrix arrays respectively. We also performed nuclear and cytoplasmic fractionation followed by Taqman low-density array and/or quantitative PCR to identify nuclear-enriched miRNAs in hemopoietic stem/progenitor cells, promyelocytes, myelocytes, granulocytes and several hemopoietic cell lines. Anti-correlation between the expression of miRNA and target pairs was used to determine putative miRNA targets.ResultsAnalyses of our array data revealed distinct clusters of differentially expressed miRNAs that are specific to promyelocytes and granulocytes. While the roles of many of these miRNAs in granulopoiesis are not currently known, anti-correlation of the expression of miRNA/mRNA target pairs identified a suite of novel target genes. Clusters of miRNAs (including members of the let-7 and miR-17-92 families) are downregulated in hemopoietic stem/progenitor cells, potentially allowing the expression of target genes known to facilitate stem cell proliferation and homeostasis. Additionally, four miRNAs (miR-709, miR-706, miR-690 and miR-467a*) were found to be enriched in the nucleus of myeloid cells and multiple hemopoietic cell lines compared to other miRNAs, which are predominantly cytoplasmic-enriched. Both miR-709 and miR-706 are nuclear-enriched throughout granulopoiesis and have putative binding sites of extensive complementarity downstream of pri-miRNAs. Nuclear enrichment of miR-467a* is specific to hemopoietic stem/progenitors and promyelocytes. These miRNAs are also nuclear-enriched in other hemopoietic cell lines, where nuclear sequestering may fine-tune the expression of cytoplasmic mRNA targets.ConclusionsOverall, we have demonstrated differentially expressed miRNAs that have not previously been associated with hemopoietic differentiation and provided further evidence of regulated nuclear-enrichment of miRNAs. Further studies into miRNA function in granulocyte development may shed light on fundamental aspects of regulatory RNA biology and the role of nuclear miRNAs.

Developmental regulation of myeloerythroid progenitor function by the LIN28B-LET-7-HMGA2 axis

“…This observation agrees with previous studies that let-7 expression increases during terminal granulocyte differentiation (Wong et al, 2014) and that disruption of Lin28 expression can affect granulopoiesis (Wang et al, 2015). These results also parallel studies from the leukemia literature wherein let-7 activity has been shown to promote myeloblast differentiation Emmrich et al, 2013;Pelosi et al, 2013).…”

Nuclear microRNAs in normal hemopoiesis and cancer

“…The functions of these nuclear miRNAs include regulation of gene and long non-coding RNA expression [37, 38, 46], controlling the biogenesis of other miRNAs [41] and fine-tuning the expression of mRNA expression in the cytoplasm [43] (Fig. 1).…”

“…Stat1 )Increased expression of transcription factor (e.g. Stat1 ) to promote granulopoiesis[43, 57]miR-690Myeloid (Lin − Sca + Kit + hemopoietic stem/progenitor cells, promyelocytes, myelocytes and granulocytes), MPRO, EL4, MEL, A20UnknownUnknown[43]miR-709Myeloid (Lin − Sca + Kit + hemopoietic stem/progenitor cells, promyelocytes, myelocytes and granulocytes), MPRO, EL4, MEL, A20UnknownUnknown[43]miR-467a*Lin − Sca + Kit + hemopoietic stem/progenitor cells, MPRO, EL4, A20UnknownUnknown[43] …”

“…Our group has recently reported the enrichment of miRNAs including miR-690, miR-706 and miR-709 in the nucleus of primary Lin − Sca + Kit + hemopoietic stem/progenitor cells, promyelocytes, myelocytes and granulocytes [43]. We and others have also found miR-690 and/or miR-709 to be nuclear-enriched in various human and mouse cell lines including MPRO, EL4, MEL, A20, L929 and HEK-293T indicating that their functions may be localized to the nucleus [41, 43].…”

miR-706 inhibits the oxidative stress-induced activation of PKCα/TAOK1 in liver fibrogenesis

“…Altogether, these results indicated that miR-706 is a hepatocyte-specific and nucleus-rich miRNA. Concomitant with our data, miR-706 was reported as a nuclear-enriched miRNA48. Since no data were available on the function of human miR-706, we next sought to locate the expression of miR-706 in human hepatocytes.…”