Oral pathogen - mediated pro-tumorigenic transformation through disruption of an Adherens Junction - associated RNAi machinery

通过破坏粘附连接相关的 RNAi 机制，口腔病原体介导促肿瘤转化

• 批准号: 10752248
• 负责人: Christina Rachel Kingsley
• 金额: $ 5.07万
• 依托单位: MEDICAL UNIVERSITY OF SOUTH CAROLINA
• 依托单位国家: 美国
• 财政年份: 2024
• 资助国家: 美国
• 起止时间: 2024-01-01 至 2026-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10752248
• 关键词: 3-Dimensional; Adherens Junction; Binding; Biological Markers; Biology; Cancer Patient; Carcinoma; Cell model; Cell-Cell Adhesion; Cells; Chad; Clustered Regularly Interspaced Short Palindromic Repeats; Coculture Techniques; Complex; Confocal Microscopy; Data; Databases; Dental; Development; Diagnosis; Disease; Doctor of Philosophy; E-Cadherin; Epithelial Cells; Epithelium; Fellowship; Fostering; Fusobacterium nucleatum; Future; Homeostasis; Immunofluorescence Immunologic; Immunoprecipitation; In Vitro; Inflammation; Intervention; Knock-out; Knowledge; Link; Malignant Neoplasms; Mediating; Messenger RNA; MicroRNAs; Microbe; Microprocessor; Molecular; Mouth Neoplasms; Oncogenes; Oncogenic; Oral; Oral cavity; Oral mucous membrane structure; Outcome; Pathologist; Pathway interactions; Patients; Periodontal Diseases; Persons; Physiological; Physiology; Protein Analysis; Publishing; Quantitative Reverse Transcriptase PCR; RNA; RNA Interference; RNA analysis; RNA-Induced Silencing Complex; Regulation; Risk; Risk Factors; Sampling; Scientist; Site; Stratified Epithelium; Survival Rate; Testing; The Cancer Genome Atlas; Therapeutic; Tissues; Training; United States; Up-Regulation; Work; cancer risk; cell behavior; cell transformation; crosslink; inflammatory marker; innovation; malignant mouth neoplasm; novel; oral bacteria; oral cavity epithelium; oral pathogen; oral tissue; oral tumorigenesis; pathogen; pathogenic bacteria; periodontopathogen; posttranscriptional; promoter; protein expression; recruit; success; successful intervention; therapeutic RNA; transcriptome sequencing; tumor progression; tumorigenic; two-dimensional

Abstract: Approximately 53,000 people in the United States are diagnosed with oral cancer yearly, and the 5- year survival rate remains poor due to late diagnosis and intervention. Recent evidence has suggested periodontal disease as a potential risk factor, and it is also associated with poor outcomes. Interestingly, periodontal disease and oral cancer share common phenomenon: disruption of epithelial barrier integrity and inflammation. More specifically, the periodontal pathogen Fusobacterium nucleatum has been associated with both barrier disruption and with oral cancer. Still, it is unclear whether disruption of the oral epithelial barrier by Fusobacterium nucleatum is a mechanistic promoter of pro-tumorigenic oral transformation, or just a consequence of it. To answer this question, we interrogate here a cellular complex that is essential for epithelial barrier integrity, called the adherens junction. More specifically, we have found that the adherens junction component PLEKHA7, which is an E-cadherin and p120 binding partner, recruits core components of the RNA interference machinery including the microprocessor, the RNA induced silencing complex, and sets of mRNAs and miRNAs to maintain epithelial homeostasis. PLEKHA7 depletion from cultured epithelial cells results in mis- localization of the RNAi machinery, decreased miRNA silencing activity, increased oncogene expression, and pro-tumorigenic cell transformation. However, such disruption of the RNAi interference machinery, particularly associated with barrier integrity, has not been studied to date in oral cancer, exposing a significant gap in knowledge. We hypothesize that PLEKHA7 recruits and regulates the RNAi machinery at oral epithelial adherens junctions to maintain homeostasis, and that this mechanism is disrupted in the presence of oral pathogens, which in turn promote expression of pro-tumorigenic and inflammatory markers. We will test our hypothesis through two Specific Aims: 1) determine whether oral epithelial adherens junctions recruit the RNAi machinery to regulate function and levels of miRNAs and of their target mRNAs, using two dimensional and three-dimensional epithelial cell models and by examining tissues from patients; 2) determine if the oral periodontal pathogen Fusobacterium nucleatum promotes upregulation of oncogenic and pro-inflammatory markers through miRNA dysregulation, using co-cultures with Fusobacterium nucleatum, immunofluorescence, protein, and RNA analyses. This project is significant, since it will identify a novel molecular mechanism of epithelial pro-tumorigenic transformation in the oral mucosa, advancing our understanding of the disease. The study is innovative, as it links pathogens, oral epithelial barrier function, and localized RNAi and miRNA regulation with oral cancer. We anticipate that the study will be impactful and contribute to future development of effective biomarkers oral cancer, which are critically needed for timely and successful intervention, as well as to the potential development of RNA-based therapeutics. Altogether, this fellowship will provide foundational training in oral epithelial biology to foster the development of the DMD/PhD trainee into a unique and critically needed academic dental scientist.

翻译后摘要：大约53,000人在美国被诊断为口腔癌每年，和5- 由于诊断和干预较晚，1年生存率仍然很低。新近证据已表明 牙周病是一个潜在的危险因素，它也与不良后果有关。有趣的是， 牙周病和口腔癌具有共同的现象：上皮屏障完整性的破坏， 炎症更具体地说，牙周病原体具核梭杆菌与 屏障破坏和口腔癌。尽管如此，目前还不清楚是否破坏口腔上皮屏障， 具核梭杆菌是促肿瘤发生口腔转化的机制启动子，或者仅仅是一种 为了回答这个问题，我们在这里询问上皮细胞所必需的细胞复合体。 屏障完整性，称为粘附连接。更具体地说，我们发现粘附连接 组分PLEKHA 7是E-钙粘蛋白和p120结合伴侣，募集RNA的核心组分 干扰机制包括微处理器、RNA诱导沉默复合物和mRNA组 和miRNAs来维持上皮细胞的稳态。从培养的上皮细胞中去除PLEKHA 7导致错配。 RNAi机制的定位，降低miRNA沉默活性，增加癌基因表达， 促肿瘤细胞转化。然而，这种RNAi干扰机制的破坏，特别是 与屏障完整性相关，迄今为止尚未在口腔癌中进行研究，暴露了在口腔癌中的显著差距。 知识我们假设PLEKHA 7在口腔上皮粘附细胞中募集并调节RNAi机制， 连接以维持体内平衡，并且这种机制在口腔病原体的存在下被破坏， 反过来促进促肿瘤发生和炎症标记物表达。我们将通过以下方式来验证我们的假设： 两个具体目的：1）确定口腔上皮粘附连接是否募集RNAi机制来调节 使用二维和三维上皮细胞模型， 细胞模型和检查患者的组织; 2）确定口腔牙周病原体梭杆菌 核质通过miRNA失调促进致癌和促炎标志物的上调， 使用与具核梭杆菌的共培养、免疫荧光、蛋白质和RNA分析。这个项目 是重要的，因为它将确定一种新的上皮促肿瘤转化的分子机制， 口腔粘膜，推进我们对疾病的理解。这项研究具有创新性，因为它将病原体、口腔 上皮屏障功能，以及口腔癌的局部RNAi和miRNA调控。我们预计 研究将是有影响，并有助于未来开发有效的生物标志物口腔癌， 及时和成功的干预，以及基于RNA的潜在发展， 治疗学总之，该奖学金将提供口腔上皮生物学的基础培训，以促进 将DMD/PhD实习生培养成独特且急需的学术牙科科学家。