Cigarette smoke induces C/EBP-β-mediated activation of miR-31 in normal human respiratory epithelia and lung cancer cells.
Cigarette smoke induces C/EBP-β-mediated activation of miR-31 in normal human respiratory epithelia and lung cancer cells.
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DOI：
10.1371/journal.pone.0013764
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发表时间：
2010-10-29
期刊：
影响因子：
3.7
通讯作者：
Schrump DS
中科院分区：
文献类型：
--
作者：
Xi S;Yang M;Tao Y;Xu H;Shan J;Inchauste S;Zhang M;Mercedes L;Hong JA;Rao M;Schrump DS
关于miRNAs促进肺癌发生的机制的信息有限。本研究旨在检测香烟烟雾冷凝物（CSC）诱导的人正常呼吸道上皮细胞和肺癌细胞中miRNAs的表达和功能。微阵列和定量RT-PCR（qRT-PCR）技术用于评估培养细胞和手术标本中的miRNA和宿主基因表达。采用软件引导分析、RNA交联免疫沉淀（CLIP）、3′ UTR荧光素酶报告基因分析、qRT-PCR、聚焦超阵列和western blot技术鉴定和确认miR-31的靶点。染色质免疫沉淀（ChIP）技术被用来评估组蛋白标记和转录因子内的LOC 554202启动子。细胞计数和异种移植实验用于评估miR-31对肺癌细胞增殖和致瘤性的影响。CSC显著增加了正常呼吸道上皮细胞和肺癌细胞中miR-31的表达并激活了LOC 554202;停止CSC暴露后，miR-31和LOC 554202的表达持续存在。肺癌组织中miR-31和LOC 554202的表达水平显著高于癌旁正常肺组织。CLIP和报告基因分析证明了miR-31与Dickkopf-1（Dkk-1）和DACT-3的直接相互作用。miR-31的过表达显著降低了正常呼吸道上皮和肺癌细胞中Dkk-1和DACT 3的表达水平。miR-31的敲低增加了Dkk-1和DACT-3水平，并消除了CSC介导的Dkk-1和DACT-3表达的降低。此外，miR-31的过表达减少了SFRP 1、SFRP 4和WIF-1，并增加了Wnt-5a的表达。CSC增加L0 C554202启动子内的H3 K4 Me 3、H3 K9/14 Ac和C/EBP-β水平。C/EBP-β的敲低消除了CSC介导的L0 C554202活化。miR-31的过表达显著增强了肺癌细胞的增殖和致瘤性; miR-31的敲低抑制了这些细胞的生长。香烟烟雾诱导miR-31的表达，其靶向正常呼吸道上皮细胞和肺癌细胞中癌症干细胞信号传导的几种拮抗剂。miR-31在人肺癌发生过程中起着oncomir的作用。
Limited information is available regarding mechanisms by which miRNAs contribute to pulmonary carcinogenesis. The present study was undertaken to examine expression and function of miRNAs induced by cigarette smoke condensate (CSC) in normal human respiratory epithelia and lung cancer cells. Micro-array and quantitative RT-PCR (qRT-PCR) techniques were used to assess miRNA and host gene expression in cultured cells, and surgical specimens. Software-guided analysis, RNA cross-link immunoprecipitation (CLIP), 3′ UTR luciferase reporter assays, qRT-PCR, focused super-arrays and western blot techniques were used to identify and confirm targets of miR-31. Chromatin immunoprecipitation (ChIP) techniques were used to evaluate histone marks and transcription factors within the LOC554202 promoter. Cell count and xenograft experiments were used to assess effects of miR-31 on proliferation and tumorigenicity of lung cancer cells. CSC significantly increased miR-31 expression and activated LOC554202 in normal respiratory epithelia and lung cancer cells; miR-31 and LOC554202 expression persisted following discontinuation of CSC exposure. miR-31 and LOC554202 expression levels were significantly elevated in lung cancer specimens relative to adjacent normal lung tissues. CLIP and reporter assays demonstrated direct interaction of miR-31 with Dickkopf-1 (Dkk-1) and DACT-3. Over-expression of miR-31 markedly diminished Dkk-1 and DACT3 expression levels in normal respiratory epithelia and lung cancer cells. Knock-down of miR-31 increased Dkk-1 and DACT3 levels, and abrogated CSC-mediated decreases in Dkk-1 and DACT-3 expression. Furthermore, over-expression of miR-31 diminished SFRP1, SFRP4, and WIF-1, and increased Wnt-5a expression. CSC increased H3K4Me3, H3K9/14Ac and C/EBP-β levels within the LOC554202 promoter. Knock-down of C/EBP-β abrogated CSC-mediated activation of LOC554202. Over-expression of miR-31 significantly enhanced proliferation and tumorigenicity of lung cancer cells; knock-down of miR-31 inhibited growth of these cells. Cigarette smoke induces expression of miR-31 targeting several antagonists of cancer stem cell signaling in normal respiratory epithelia and lung cancer cells. miR-31 functions as an oncomir during human pulmonary carcinogenesis.
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影响因子：
3.7
作者：
Corcoran DL;Pandit KV;Gordon B;Bhattacharjee A;Kaminski N;Benos PV
通讯作者：
Benos PV
影响因子：
4.7
作者：
Izzotti, Alberto;Larghero, Patrizia;De Flora, Silvio
通讯作者：
De Flora, Silvio
影响因子：
254.7
作者：
Glynn, Thomas;Seffrin, John R.;Ross, Hana
通讯作者：
Ross, Hana
影响因子：
11.2
作者：
Creighton CJ;Fountain MD;Yu Z;Nagaraja AK;Zhu H;Khan M;Olokpa E;Zariff A;Gunaratne PH;Matzuk MM;Anderson ML
通讯作者：
Anderson ML
影响因子：
3.7
作者：
Gluschnaider U;Hidas G;Cojocaru G;Yutkin V;Ben-Neriah Y;Pikarsky E
通讯作者：
Pikarsky E