Zini Roberta, Rossi Chiara, Norfo Ruggiero, Pennucci Valentina, Barbieri Greta, Ruberti Samantha, Rontauroli Sebastiano, Salati Simona, Bianchi Elisa, Manfredini Rossella
1 Centre for Regenerative Medicine "Stefano Ferrari," Department of Life Sciences, University of Modena and Reggio Emilia , Modena, Italy .
2 Haematopoietic Stem Cell Biology Laboratory, Weatherall Institute of Molecular Medicine, University of Oxford , Oxford, United Kingdom .
Stem Cells Dev. 2016 Oct 1;25(19):1433-43. doi: 10.1089/scd.2016.0150. Epub 2016 Sep 7.
microRNAs are key regulators of gene expression that control stem cell fate by posttranscriptional downregulation of hundreds of target genes through seed pairing in their 3' untranslated region. In fact, miRNAs tightly regulate fundamental stem cell processes, like self-renewal, proliferation, and differentiation; therefore, miRNA deregulation may contribute to the development of solid tumors and hematological malignancies. miR-382-5p has been found to be upregulated in patients with myeloid neoplasms, but its role in normal hematopoiesis is still unknown. In this study, we demonstrated that miR-382-5p overexpression in CD34(+) hematopoietic stem/progenitor cells (HSPCs) leads to a significant decrease of megakaryocyte precursors coupled to increase of granulocyte ones. Furthermore, by means of a computational analysis using different prediction algorithms, we identified several putative mRNA targets of miR-382-5p that are downregulated upon miRNA overexpression (ie, FLI1, GATA2, MAF, MXD1, RUNX1, and SGK1). Among these, we validated MXD1 as real target of miR-382-5p by luciferase reporter assay. Finally, we showed that MXD1 knockdown mimics the effects of miR-382-5p overexpression on granulocyte and megakaryocyte differentiation of CD34(+) cells. Overall, our results demonstrated that miR-382-5p expression favors the expansion of granulocyte lineage and impairs megakaryocyte commitment through MXD1 downregulation. Therefore, our data showed for the first time that the miR-382-5p/MXD1 axis plays a critical role in myelopoiesis by affecting the lineage choice of CD34(+) HSPCs.
微小RNA是基因表达的关键调节因子，通过其3'非翻译区的种子配对在转录后下调数百个靶基因，从而控制干细胞命运。事实上，微小RNA紧密调节干细胞的基本过程，如自我更新、增殖和分化；因此，微小RNA失调可能导致实体瘤和血液系统恶性肿瘤的发生。已发现miR-382-5p在髓系肿瘤患者中上调，但其在正常造血中的作用仍不清楚。在本研究中，我们证明CD34(+)造血干/祖细胞（HSPCs）中miR-382-5p的过表达导致巨核细胞前体显著减少，同时粒细胞前体增加。此外，通过使用不同预测算法的计算分析，我们鉴定了miR-382-5p的几个假定mRNA靶标，这些靶标在miRNA过表达时被下调（即FLI1、GATA2、MAF、MXD1、RUNX1和SGK1）。其中，我们通过荧光素酶报告基因检测验证了MXD1是miR-382-5p的真正靶标。最后，我们表明MXD1敲低模拟了miR-382-5p过表达对CD34(+)细胞粒细胞和巨核细胞分化的影响。总体而言，我们的结果表明miR-382-5p表达有利于粒细胞谱系的扩增，并通过下调MXD1损害巨核细胞的定向分化。因此，我们的数据首次表明miR-382-5p/MXD1轴通过影响CD34(+) HSPCs的谱系选择在髓系造血中起关键作用。
