Differential regulation of MMPs by E2F1, Sp1 and NF-kappa B controls the small cell lung cancer invasive phenotype.
Differential regulation of MMPs by E2F1, Sp1 and NF-kappa B controls the small cell lung cancer invasive phenotype.
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E2F1、Sp1 和 NF-kappa B 对 MMP 的差异调节控制着小细胞肺癌的侵袭表型。
DOI：
10.1186/1471-2407-14-276
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发表时间：
2014-04-22
期刊：
影响因子：
3.8
通讯作者：
Yuan H
中科院分区：
文献类型：
--
作者：
Li Z;Guo Y;Jiang H;Zhang T;Jin C;Young CY;Yuan H
研究背景E2F1转录因子在细胞增殖、凋亡、侵袭和转移等多种细胞过程的调节中发挥着至关重要的作用。 E2F1在小细胞肺癌（SCLC）中过度表达，早期广泛转移是SCLC最重要的特征。在本研究中，我们通过调节基质金属蛋白酶（MMPs）的表达来探讨E2F1在SCLC侵袭和转移过程中的参与。方法采用免疫组织化学方法检测中国汉族人群SCLC样本中E2F1和MMPs的表达。通过 Transwell 和伤口愈合实验观察 E2F1 对侵袭和转移的影响，并通过特异性 siRNA 耗竭 E2F1。通过染色质免疫沉淀（ChIP）测序鉴定E2F1调控的靶基因，并通过实时PCR和蛋白质印迹检测靶基因的表达。采用双荧光素酶报告系统分析E2F1与MMPs之间的调控关系。结果E2F1是中国汉族人群SCLC中高表达的独立的不良预后因子。通过特异性 siRNA 敲低 E2F1 导致 SCLC 中迁移和侵袭的下调。 SCLC中MMP-9和-16的表达高于其他MMP，且沉默E2F1后其表达降低最显着。 ChIP 序列和基于启动子的荧光素酶分析表明，E2F1 通过启动子中的 E2F1 结合基序直接控制 MMP-16 的表达。尽管预测 MMP-9 启动子中有一个 E2F1 结合位点，但荧光素酶分析表明该结合位点在功能上并不是必需的。进一步的研究表明，E2F1转录控制Sp1和p65的表达，从而增强SCLC细胞中MMP-9启动子的活性。通过 SCLC 肿瘤中的免疫组织化学染色验证了 E2F1、Sp1、p65 和 MMP-9 之间的关联。结论 E2F1 作为 MMP 的转录激活剂，通过与启动子中的 E2F1 结合序列结合直接增强 MMP 转录，或通过增强的 Sp1 和 NF-κ B 间接激活 MMP SCLC 中 E2F1 激活。
BackgroundE2F1 transcription factor plays a vital role in the regulation of diverse cellular processes including cell proliferation, apoptosis, invasion and metastasis. E2F1 overexpression has been demonstrated in small cell lung cancer (SCLC), and extensive metastasis in early phase is the most important feature of SCLC. In this study, we investigated the involvement of E2F1 in the process of invasion and metastasis in SCLC by regulating the expression of matrix metalloproteinases (MMPs).MethodsImmunohistochemistry was performed to evaluate the expression of E2F1 and MMPs in SCLC samples in a Chinese Han population. The impact of E2F1 on invasion and metastasis was observed by transwell and wound healing experiments with depletion of E2F1 by specific siRNA. The target genes regulated by E2F1 were identified by chromatin immunoprecipitation (ChIP)-to-sequence, and the expressions of target genes were detected by real time PCR and western blotting. The dual luciferase reporter system was performed to analyze the regulatory relationship between E2F1 and MMPs.ResultsE2F1 is an independent and adverse prognosis factor that is highly expressed in SCLC in a Chinese Han population. Knockdown of E2F1 by specific siRNA resulted in the downregulation of migration and invasion in SCLC. The expressions of MMP-9 and −16 in SCLC were higher than other MMPs, and their expressions were most significantly reduced after silencing E2F1. ChIP-to-sequence and promoter-based luciferase analysis demonstrated that E2F1 directly controlled MMP-16 expression via an E2F1 binding motif in the promoter. Although one E2F1 binding site was predicted in the MMP-9 promoter, luciferase analysis indicated that this binding site was not functionally required. Further study demonstrated that E2F1 transcriptionally controlled the expression of Sp1 and p65, which in turn enhanced the MMP-9 promoter activity in SCLC cells. The associations between E2F1, Sp1, p65, and MMP-9 were validated by immunohistochemistry staining in SCLC tumors.ConclusionsE2F1 acts as a transcriptional activator for MMPs and directly enhances MMP transcription by binding to E2F1 binding sequences in the promoter, or indirectly activates MMPs through enhanced Sp1 and NF-kappa B as a consequence of E2F1 activation in SCLC.
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作者：
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