AP-2α Induces Epigenetic Silencing of Tumor Suppressive Genes and Microsatellite Instability in Head and Neck Squamous Cell Carcinoma

BackgroundActivator protein 2 alpha (AP-2α) is involved in a variety of physiological processes. Increased AP-2α expression correlates with progression in various squamous cell carcinomas, and a recent publication found AP-2α to be overexpressed in ∼70% of Head and Neck Squamous Cell Carcinoma (HNSCC) patient samples. It was found to repress transcription of the tumor suppressor gene C/CAAT Enhancer Binding Protein alpha (C/EBPα), and its binding site correlated with upstream methylation of the C/EBPα promoter. Therefore, we investigated the potential for AP-2α to target methylation to additional genes that would be relevant to HNSCC pathogenesis.
Principal FindingsStable downregulation of AP-2α stable by shRNA in HNSCC cell lines correlated with decreased methylation of its target genes' regulatory regions. Furthermore, methylation of MLH1 in HNSCC with and without AP-2α downregulation revealed a correlation with microsatellite instability (MSI). ChIP analysis was used to confirm binding of AP-2α and HDAC1/2 to the targets. The effects of HDAC inhibition was assessed using Trichostatin A in a HNSCC cell line, which revealed that AP-2α targets methylation through HDAC recruitment.
ConclusionsThese findings are significant because they suggest AP-2α plays a role not only in epigenetic silencing, but also in genomic instability. This intensifies the potential level of regulation AP-2α has through transcriptional regulation. Furthermore, these findings have the potential to revolutionize the field of HNSCC therapy, and more generally the field of epigenetic therapy, by targeting a single gene that is involved in the malignant transformation via disrupting DNA repair and cell cycle control.

背景
激活蛋白2α（Activator Protein 2 alpha，AP-2α）参与多种生理进程。AP-2α表达上调与多种鳞状细胞癌的进展相关，近期一项研究发现，约70%的头颈部鳞状细胞癌（Head and Neck Squamous Cell Carcinoma，HNSCC）患者样本中AP-2α呈过表达状态。研究证实，AP-2α可抑制抑癌基因C/CAAT增强子结合蛋白α（C/CAAT Enhancer Binding Protein alpha，C/EBPα）的转录，且其结合位点与C/EBPα启动子的上游甲基化水平相关。因此，本研究旨在探究AP-2α将甲基化靶向调控其他与HNSCC发病机制相关基因的潜在可能。
主要研究结果
通过短发夹RNA（short hairpin RNA，shRNA）稳定下调HNSCC细胞系中的AP-2α表达后，其靶基因调控区域的甲基化水平显著降低。此外，对AP-2α下调前后的HNSCC细胞中MLH1的甲基化水平分析显示，其与微卫星不稳定性（Microsatellite Instability，MSI）存在相关性。采用染色质免疫沉淀（Chromatin Immunoprecipitation，ChIP）实验验证了AP-2α及组蛋白去乙酰化酶1/2（Histone Deacetylase 1/2，HDAC1/2）与靶位点的结合。使用曲古抑菌素A（Trichostatin A，TSA）处理HNSCC细胞系以评估HDAC抑制剂的作用效果，结果表明AP-2α通过招募HDAC实现靶基因的甲基化调控。
结论
本研究结果具有重要学术价值，因其表明AP-2α不仅参与表观遗传沉默，还可影响基因组不稳定性。这进一步强化了AP-2α通过转录调控实现的多层级调控作用。此外，本研究发现有望革新HNSCC治疗领域，乃至更广范围的表观遗传治疗领域——通过靶向单一通过破坏DNA修复与细胞周期调控参与恶性转化的基因。