Yuan Xin, Liu Pengcong, Xu Lei, Liang Liqing, Dong Qian, Fan Tao, Yue Wen, Qu Mingyi, Pei Xuetao, Xie Xiaoyan
Stem Cell and Regenerative Medicine Lab, Beijing Institute of Radiation Medicine, No. 27 Taiping Road, Haidian District, Beijing, 100850, China.
Thromb J. 2024 Aug 9;22(1):74. doi: 10.1186/s12959-024-00615-6.
Proper control of the lineage bias of megakaryocytic and erythroid progenitor cells (MEPs) is of significant importance, the disorder of which will lead to abnormalities in the number and function of platelets and erythrocytes. Unfortunately, the signaling pathways regulating MEP differentiation largely remain to be elucidated. This study aimed to analyze the role and the underlying molecular mechanism of miR-1915-3p in megakaryocytic and erythroid differentiation.
We utilized miRNA mimics and miRNA sponge to alter the expression of miR-1915-3p in megakaryocytic and/or erythroid potential cells; siRNA and overexpression plasmid to change the expression of SOCS4, a potential target of miR-1915-3p. The expression of relevant surface markers was detected by flow cytometry. We scanned for miR-1915-3p target genes by mRNA expression profiling and bioinformatic analysis, and confirmed the targeting by dual-luciferase reporter assay, western blot and gain- and lost-of-function studies. One-way ANOVA and t-test were used to analyze the statistical significance.
In this study, overexpression or knockdown of miR-1915-3p inhibited or promoted erythroid differentiation, respectively. Accordingly, we scanned for miR-1915-3p target genes and confirmed that SOCS4 is one of the direct targets of miR-1915-3p. An attentive examination of the endogenous expression of SOCS4 during megakaryocytic and erythroid differentiation suggested the involvement of SOCS4 in erythroid/megakaryocytic lineage determination. SOCS4 knockdown lessened erythroid surface markers expression, as well as improved megakaryocytic differentiation, similar to the effects of miR-1915-3p overexpression. While SOCS4 overexpression resulted in reversed effects. SOCS4 overexpression in miR-1915-3p upregulated cells rescued the effect of miR-1915-3p.
miR-1915-3p acts as a negative regulator of erythropoiesis, and positively in thrombopoiesis. SOCS4 is one of the key mediators of miR-1915-3p during the differentiation of MEPs.
正确控制巨核细胞和红系祖细胞（MEP）的谱系偏向至关重要，其紊乱会导致血小板和红细胞数量及功能异常。遗憾的是，调节MEP分化的信号通路很大程度上仍有待阐明。本研究旨在分析miR-1915-3p在巨核细胞和红系分化中的作用及潜在分子机制。
我们利用miRNA模拟物和miRNA海绵来改变巨核细胞和/或红系潜能细胞中miR-1915-3p的表达；利用小干扰RNA（siRNA）和过表达质粒来改变miR-1915-3p的潜在靶标SOCS4的表达。通过流式细胞术检测相关表面标志物的表达。我们通过mRNA表达谱分析和生物信息学分析筛选miR-1915-3p靶基因，并通过双荧光素酶报告基因检测、蛋白质免疫印迹以及功能获得和缺失研究来证实靶向作用。采用单因素方差分析和t检验分析统计学意义。
在本研究中，miR-1915-3p的过表达或敲低分别抑制或促进红系分化。相应地，我们筛选了miR-1915-3p靶基因，并证实SOCS4是miR-1915-3p的直接靶标之一。对巨核细胞和红系分化过程中SOCS4内源性表达的仔细检查表明，SOCS4参与红系/巨核细胞谱系的决定。敲低SOCS4可降低红系表面标志物的表达，并改善巨核细胞分化，这与过表达miR-1915-3p的效果相似。而SOCS4过表达则产生相反的效果。在miR-1915-3p上调的细胞中过表达SOCS4可挽救miR-1915-3p的作用。
miR-1915-3p是红细胞生成的负调节因子，对血小板生成起正调节作用。SOCS4是miR-1915-3p在MEP分化过程中的关键介质之一。
