Regulation of ZEB1 by Porphyromonas gingivalis in gingival epithelial cells

牙龈上皮细胞中牙龈卟啉单胞菌对ZEB1的调节

• 批准号: 9980359
• 负责人: Zackary R Fitzsimonds
• 金额: $ 4.29万
• 依托单位: UNIVERSITY OF LOUISVILLE
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9980359
• 关键词: Amanitins; Anaerobic Bacteria; Apical; Apoptosis; Binding; Biopsy; Blood Circulation; Cancer Patient; Cell Cycle; Cell Cycle Progression; Cell Nucleus; Cells; Co-Immunoprecipitations; Complex; Data; Development; Diagnosis; E-Cadherin; E1A-associated p300 protein; Embryonic Development; Environment; Epidemiology; Epithelial; Epithelial Cells; Epithelium; Esophageal Squamous Cell Carcinoma; Family; Foundations; Future; Gene Expression; Genes; Genetic Transcription; Germ Cells; Gingiva; Homeobox; Human; Immunofluorescence Immunologic; Immunohistochemistry; Immunoprecipitation; Intercept; Invaded; Knowledge; Laboratories; Lead; Literature; Luciferases; MADH2 gene; MADH3 gene; MADH4 gene; MADH6 gene; MADH7 gene; MMP9 gene; Malignant Neoplasms; Measures; Mediating; Mesenchymal; MicroRNAs; Migration Assay; Molecular; N-Cadherin; Neoplasm Metastasis; Nuclear; Oncogenic; Oral; Oral cavity; Pathway interactions; Patients; Periodontal Diseases; Periodontitis; Phenotype; Phosphorylation; Plasmids; Play; Porifera; Porphyromonas gingivalis; Primary carcinoma of the liver cells; Process; Property; Quantitative Reverse Transcriptase PCR; RNA; RNA Synthesis Inhibitors; Ramp; Regulation; Role; Sampling; Signal Pathway; Signal Transduction; Speed; Survival Rate; TGF Beta Signaling Pathway; Testing; Tight Junctions; Time; Tongue Neoplasms; Transcription Coactivator; Transforming Growth Factor beta Receptors; Translating; Untranslated RNA; Up-Regulation; Western Blotting; Zinc Fingers; arm; cancer cell; cancer diagnosis; differential expression; dysbiosis; epithelial to mesenchymal transition; expression vector; genetic corepressor; host-microbe interactions; human tissue; knock-down; mRNA Expression; mRNA Stability; malignant mouth neoplasm; mouth squamous cell carcinoma; new therapeutic target; novel; novel diagnostics; novel therapeutics; outcome forecast; overexpression; prevent; promoter; recruit; response; scaffold; synergism; transcription factor; tumor

Project Summary Oral cancer is the 11th most frequently diagnosed cancer in the world, and due to its late diagnosis, it has an approximately 50% survival rate after 5 years. Metastasis is meditated through Epithelial to Mesenchymal Transition (EMT), which is the process by which epithelial cells lose their tight junctions and apical/basal polarity and shift to a more invasive mesenchymal phenotype giving the cancer cells systemic access through invasion of the bloodstream. EMT is mediated through several transcriptional factors, however we will focus on ZEB1. Porphyromonas gingivalis is an oral obligate anaerobe that when present shifts the host/microbe relationship from synergy to a more dysbiotic environment, which contributes to periodontal disease. P. gingivalis has been shown to invade gingival epithelial cells, and once inside the cells can inhibit apoptosis, ramp up the cell cycle, and more recently by our lab has been shown to upregulate ZEB1. Although we are beginning to understand the potential oncogenic role of P. gingivalis, we do not currently know how ZEB1 is regulated by P. gingivalis in gingival epithelial cells. Our preliminary data suggests that this regulation is occurring through SMAD signaling regulation and through the long non-coding RNA ZEB1-AS1. SMAD signaling is a branch of the TGF-b signaling pathway, which during embryonic development is responsible for producing mesenchymal cells from the initial epithelium. SMAD signaling occurs by phosphorylation of SMAD2/3 by the TGF-b receptor, followed by the formation of a transcriptional regulatory trimer by SMAD2/3 and SMAD4, and finally nuclear localization and upregulation of ZEB1. Additionally, ZEB1 can bind to SMAD2/3 and SMAD4 to form a ZEB1/SMAD trimer to regulate ZEB1 transcription. We have found that P. gingivalis upregulates the regulatory SMAD3, which suggests P. gingivalis is able to intercept this pathway to upregulate ZEB1. We have also found that P. gingivalis upregulates ZEB1-AS1, which has been shown in hepatocarcinoma and esophageal squamous cell carcinoma to directly upregulate ZEB1, and our preliminary data supports this as knocking down ZEB1-AS1 leads to abrogation of ZEB1 regulation by P. gingivalis in gingival epithelial cells. We will investigate mechanisms of regulation by ZEB1-AS1 including direct promotion of gene expression in the nucleus through binding to P300, and/or control of ZEB1 mRNA stability. EMT regulation is a complex process that involves integration of several host signaling pathways, so we also suspect that ZEB1-AS1 will be integrated with SMAD signaling by potentially acting as a scaffold for SMAD3. Developing a better understanding of P. gingivalis mediated EMT will lay the foundation for future studies to target these regulatory mechanisms for diagnosis and treatment, and ultimately reduce the 50% chance of survival current patients have 5 years after diagnosis.

项目摘要 口腔癌是世界上第11位最常被诊断的癌症，由于诊断较晚，它的发病率很高。 5年后的存活率约为50%。转移是通过上皮到间质介导的 过渡（EMT），这是上皮细胞失去其紧密连接和顶端/基底极性的过程 并转变为更具侵袭性的间充质表型， 血液循环。EMT是通过几个转录因子介导的，但我们将重点放在ZEB 1。 牙龈卟啉单胞菌是一种口腔专性厌氧菌，当存在时，改变宿主/微生物关系 从协同作用到更加生态失调的环境，这有助于牙周病。牙龈卟啉单胞菌一直是 显示侵入牙龈上皮细胞，并且一旦进入细胞内就可以抑制细胞凋亡，加速细胞周期， 最近我们的实验室发现它能上调ZEB 1。虽然我们开始了解 牙龈卟啉单胞菌的潜在致癌作用，我们目前还不知道ZEB 1是如何被牙龈卟啉单胞菌调节的， 牙龈上皮细胞我们的初步数据表明，这种调节是通过SMAD信号发生的， 调控和通过长的非编码RNA ZEB 1-AS 1。SMAD信号是TGF-β信号的一个分支， 途径，在胚胎发育过程中负责从最初的间充质细胞产生 上皮SMAD信号传导通过TGF-β受体磷酸化SMAD 2/3，然后通过TGF-β受体磷酸化SMAD 2/3而发生。 通过SMAD 2/3和SMAD 4形成转录调节三聚体，最后是核定位， ZEB 1的上调。此外，ZEB 1可以结合SMAD 2/3和SMAD 4以形成ZEB 1/SMAD三聚体， 调节ZEB 1转录。我们发现牙龈卟啉单胞菌上调SMAD 3的调节，这表明 P. Gingivalis能够阻断该途径以上调ZEB 1。我们还发现牙龈卟啉单胞菌 上调ZEB 1-AS 1，在肝癌和食管鳞状细胞癌中显示 直接上调ZEB 1，我们的初步数据支持这一点，因为敲低ZEB 1-AS 1会导致 牙龈卟啉单胞菌对牙龈上皮细胞中ZEB 1调节的消除。我们将研究 ZEB 1-AS 1的调节包括通过与P300结合直接促进细胞核中的基因表达， 和/或控制ZEB 1 mRNA的稳定性。EMT监管是一个复杂的过程，涉及几个方面的整合。 因此，我们也怀疑ZEB 1-AS 1将通过潜在的途径与SMAD信号转导整合， 作为SMAD 3的支架。更好地了解牙龈卟啉单胞菌介导的EMT将为我们提供 为未来的研究奠定基础，以这些调控机制为目标进行诊断和治疗，并最终 减少了50%的生存机会，目前的患者有5年后诊断。