MicroRNA transcriptome analysis identifies miR-365 as a novel negative regulator of cell proliferation in Zmpste24-deficient mouse embryonic fibroblasts.
MicroRNA transcriptome analysis identifies miR-365 as a novel negative regulator of cell proliferation in Zmpste24-deficient mouse embryonic fibroblasts.
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DOI：
10.1016/j.mrfmmm.2015.04.010
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发表时间：
2015-07
期刊：
影响因子：
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通讯作者：
Suh Y
中科院分区：
文献类型：
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作者：
Xiong XD;Jung HJ;Gombar S;Park JY;Zhang CL;Zheng H;Ruan J;Li JB;Kaeberlein M;Kennedy BK;Zhou Z;Liu X;Suh Y
Zmpste 24是一种金属蛋白酶，负责前层蛋白A翻译后加工和切割成成熟层蛋白A。Zmpste 24-/-小鼠表现出与表达早老蛋白的小鼠重叠的一系列早老样表型，早老蛋白是与Hutchinson-Gilford早老综合征（HGPS）相关的核纤层蛋白A的改变形式。越来越多的证据表明，miRNAs参与了正常衰老过程的调控，但其在早衰症中的作用尚不清楚。在这里，我们报告了小鼠胚胎成纤维细胞（MEFs）建立从野生型（WT）和Zmpste 24-/- progeroid小鼠使用大规模平行测序技术的miRNA转录组。利用来自WT MEFs的19.5 ×106个读段和来自Zmpste 24-/- MEFs的16.5 × 106个读段的数据，我们发现了在MEFs中表达的总共306个已知的miRNA，其读段计数的动态范围从10到超过100万。与WT MEF相比，在Zmpste 24-/-MEF中发现总共8种miRNA显著下调，仅2种miRNA上调。功能研究表明，miR-365，Zmpste 24-/- MEFs中显著下调的miRNA，调节MEFs中的细胞生长表型。在Zmpste 24-/- MEFs中过表达miR-365增加细胞增殖并降低SA-β-gal阳性细胞的百分比，而抑制miR-365功能导致WT MEFs中SA-β-gal阳性细胞的增加。此外，我们确定了小GTP酶Ras超家族成员Rasd 1作为miR-365的功能性靶点。虽然miR-365的表达抑制了Rasd 1 3′UTR的报告酶活性，但这种作用在假定的3′UTR靶位点突变时丧失。一致地，发现miR-365的表达水平与内源性Rasd 1水平负相关。这些发现表明，miR-365在Zmpste 24-/- MEFs中下调，并作为Rasd 1的一种新的负调节因子。我们全面的miRNA数据为研究MEFs中的基因调控网络提供了资源。
Zmpste24 is a metalloproteinase responsible for the posttranslational processing and cleavage of prelamin A into mature laminA. Zmpste24-/- mice display a range of progeroid phenotypes overlapping with mice expressing progerin, an altered version of lamin A associated with Hutchinson-Gilford progeria syndrome (HGPS). Increasing evidence has demonstrated that miRNAs contribute to the regulation of normal aging process, but their roles in progeroid disorders remain poorly understood. Here we report the miRNA transcriptomes of mouse embryonic fibroblasts (MEFs) established from wild type (WT) and Zmpste24-/- progeroid mice using a massively parallel sequencing technology. With data from 19.5 ×106 reads from WT MEFs and 16.5 × 106 reads from Zmpste24-/- MEFs, we discovered a total of 306 known miRNAs expressed in MEFs with a wide dynamic range of read counts ranging from 10 to over 1 million. A total of 8 miRNAs were found to be significantly down-regulated, with only 2 miRNAs upregulated, in Zmpste24-/- MEFs as compared to WT MEFs. Functional studies revealed that miR-365, a significantly down-regulated miRNA in Zmpste24-/- MEFs, modulates cellular growth phenotypes in MEFs. Overexpression of miR-365 in Zmpste24-/- MEFs increased cellular proliferation and decreased the percentage of SA-β-gal-positive cells, while inhibition of miR-365 function led to an increase of SA-β-gal-positive cells in WT MEFs. Furthermore, we identified Rasd1, a member of the Ras superfamily of small GTPases, as a functional target of miR-365. While expression of miR-365 suppressed Rasd1 3′UTR luciferase-reporter activity, this effect was lost with mutations in the putative 3′UTR target-site. Consistently, expression levels of miR-365 were found to inversely correlate with endogenous Rasd1 levels. These findings suggest that miR-365 is down-regulated in Zmpste24-/- MEFs and acts as a novel negative regulator of Rasd1. Our comprehensive miRNA data provide a resource to study gene regulatory networks in MEFs.