Di Sanzo M, Chirillo R, Aversa I, Biamonte F, Santamaria G, Giovannone E D, Faniello M C, Cuda G, Costanzo F
Centro di Ricerca di Biochimica e Biologia Molecolare Avanzata, Dipartimento di Medicina Sperimentale e Clinica, Università degli Studi "Magna Græcia", Catanzaro, Italy; Dipartimento di Medicina Sperimentale e Clinica, Università degli Studi "Magna Græcia", Catanzaro, Italy.
Centro Interdipartimentale di Servizi e Ricerca, Università degli Studi "Magna Græcia", Catanzaro, Italy; Dipartimento di Medicina Sperimentale e Clinica, Università degli Studi "Magna Græcia", Catanzaro, Italy.
Gene. 2018 May 30;657:92-99. doi: 10.1016/j.gene.2018.03.027. Epub 2018 Mar 12.
The heavy subunit of the iron storage protein ferritin (FHC) is essential for the intracellular iron metabolism and, at the same time, it represents a central hub of iron-independent pathways, such as cell proliferation, angiogenesis, p53 regulation, chemokine signalling, stem cell expansion, miRNAs expression. In this work we have explored the ability of FHC to modulate gene expression in K562 cells, through the up-regulation of the lncRNA H19 and its cognate miR-675.
Targeted silencing of FHC was performed by lentiviral-driven shRNA strategy. FHC reconstitution was obtained by full length FHC cDNA transfection with Lipofectamine 2000. ROS amounts were determined with the redox-sensitive probe H2DCFDA. H19, miR-675, miR-107, Twist1, ID3, EPHB6, GNS, ANK1 and SMAD6 mRNA amounts were quantified by Taqman assay and qPCR analysis.
FHC silencing in K562 cells modulates gene expression through the up-regulation of the lncRNA H19 and its cognate miR-675. Experimental findings demonstrate that the molecular mechanism underlying this phenomenon is represented by an FHC knock-down-triggered increase in reactive oxygen species (ROS) production.
In this paper we uncover a so far not described function of the ferritin heavy subunit in the control of lncRNA pathways.
铁储存蛋白铁蛋白重链（FHC）的重亚基对细胞内铁代谢至关重要，同时，它也是铁非依赖途径的核心枢纽，如细胞增殖、血管生成、p53调控、趋化因子信号传导、干细胞扩增、微小RNA（miRNA）表达。在本研究中，我们通过上调长链非编码RNA（lncRNA）H19及其同源物miR-675，探索了FHC调节K562细胞基因表达的能力。
采用慢病毒驱动的短发夹RNA（shRNA）策略对FHC进行靶向沉默。通过用脂质体2000转染全长FHC cDNA实现FHC的重组。用氧化还原敏感探针H2DCFDA测定活性氧（ROS）含量。通过Taqman分析和定量聚合酶链反应（qPCR）分析对H19、miR-675、miR-107、Twist1、ID3、EPHB6、GNS、ANK1和SMAD6的mRNA含量进行定量。
K562细胞中的FHC沉默通过上调lncRNA H19及其同源物miR-675来调节基因表达。实验结果表明，这一现象的分子机制表现为FHC敲低引发的活性氧生成增加。
在本文中，我们揭示了铁蛋白重亚基在lncRNA途径控制中迄今未被描述的功能。
