Biamonte Flavia, Zolea Fabiana, Bisognin Andrea, Di Sanzo Maddalena, Saccoman Claudia, Scumaci Domenica, Aversa Ilenia, Panebianco Mariafranca, Faniello Maria Concetta, Bortoluzzi Stefania, Cuda Giovanni, Costanzo Francesco
Department of Experimental and Clinical Medicine, Magna Græcia University of Catanzaro, Salvatore Venuta Campus, Viale Europa, 88100, Catanzaro, Italy.
Department of Biology, University of Padua, Via G. Colombo 3, 35131, Padua, Italy.
PLoS One. 2015 Mar 27;10(3):e0122105. doi: 10.1371/journal.pone.0122105. eCollection 2015.
In a previous study, we showed that the silencing of the heavy subunit (FHC) offerritin, the central iron storage molecule in the cell, is accompanied by a modification in global gene expression. In this work, we explored whether different FHC amounts might modulate miRNA expression levels in K562 cells and studied the impact of miRNAs in gene expression profile modifications. To this aim, we performed a miRNA-mRNA integrative analysis in K562 silenced for FHC (K562shFHC) comparing it with K562 transduced with scrambled RNA (K562shRNA). Four miRNAs, namely hsa-let-7g, hsa-let-7f, hsa-let-7i and hsa-miR-125b, were significantly up-regulated in silenced cells. The remarkable down-regulation of these miRNAs, following FHC expression rescue, supports a specific relation between FHC silencing and miRNA-modulation. The integration of target predictions with miRNA and gene expression profiles led to the identification of a regulatory network which includes the miRNAs up-regulated by FHC silencing, as well as91 down-regulated putative target genes. These genes were further classified in 9 networks; the highest scoring network, "Cell Death and Survival, Hematological System Development and Function, Hematopoiesis", is composed by 18 focus molecules including RAF1 and ERK1/2. We confirmed that, following FHC silencing, ERK1/2 phosphorylation is severely impaired and that RAF1 mRNA is significantly down-regulated. Taken all together, our data indicate that, in our experimental model, FHC silencing may affect RAF1/pERK1/2 levels through the modulation of a specific set of miRNAs and add new insights in to the relationship among iron homeostasis and miRNAs.
在先前的一项研究中，我们发现，细胞中核心铁储存分子铁蛋白重链亚基（FHC）的沉默伴随着整体基因表达的改变。在这项研究中，我们探究了不同FHC含量是否会调节K562细胞中的miRNA表达水平，并研究了miRNA对基因表达谱改变的影响。为此，我们对FHC沉默的K562细胞（K562shFHC）进行了miRNA-mRNA整合分析，并将其与转导了乱序RNA的K562细胞（K562shRNA）进行比较。四种miRNA，即hsa-let-7g、hsa-let-7f、hsa-let-7i和hsa-miR-125b，在沉默细胞中显著上调。FHC表达恢复后，这些miRNA显著下调，这支持了FHC沉默与miRNA调节之间的特定关系。将靶标预测与miRNA和基因表达谱进行整合，从而鉴定出一个调控网络，其中包括因FHC沉默而上调的miRNA，以及91个下调的假定靶基因。这些基因进一步被分为9个网络；得分最高的网络“细胞死亡与存活、血液系统发育与功能、造血作用”由18个核心分子组成，包括RAF1和ERK1/2。我们证实，FHC沉默后，ERK1/2磷酸化受到严重损害，且RAF1 mRNA显著下调。综上所述，我们的数据表明，在我们的实验模型中，FHC沉默可能通过调节一组特定的miRNA来影响RAF1/pERK1/2水平，并为铁稳态与miRNA之间的关系提供了新的见解。
