The role of the primary cilium in insulin signaling and common disease pathogenesis

项目总结 少见的初级纤毛遗传缺陷可导致许多表型重叠的孟德尔 被称为纤毛病的综合征。有趣的是，许多纤毛病患者表现出血糖受损的迹象。 动态平衡，越来越多的证据表明，胰岛素受体必须被特异性地输送到 纤毛启动胰岛素依赖信号级联反应。然而，我们还不知道纤毛基因中的变异是否 更普遍地增加个人患糖尿病的风险，我们也不了解睫状毛 基因可能影响胰岛素受体信号从而导致病理改变。这项提议的目标是填补我们的这一空白 使用互补的生物信息学和细胞生物学方法的知识。我的临床和科学专长 在纤毛生物学和人类遗传学方面，加上我令人兴奋的初步结果，严谨的研究计划，以及 优秀的导师团队，为这个项目提供了坚实的基础。在完成拟议的工作时， 特别是它的重点是在生物信息学领域和实验室管理方面为我提供额外的培训， 我将作为一名内科科学家开始我的独立学术生涯，寻求更好的 了解人类疾病中睫毛功能障碍的遗传学和生物学。我已经得到了完全的支持 ，并将从两个机构提供的无与伦比的资源和指导中受益匪浅 宾夕法尼亚大学和费城儿童医院颁发这一奖项。 我假设纤毛基因的变异通过扰乱细胞信号通路而增加患糖尿病的风险 对于正常的胰岛素信号来说是必不可少的。我在英国和宾夕法尼亚医学生物库的初步分析表明 已经确定了许多与葡萄糖和血红蛋白A1c显著相关的纤毛基因和变体 水平，并显示纤毛基因表达的显著变化，以响应胰岛素。我会进一步 用两组互补的分析来研究这些关联。具体来说，我将(1)使用生物信息学 在大型患者数据集中识别和表征纤毛基因和通路的方法 增加2型糖尿病表型的风险，以及(2)在疾病的细胞模型中验证这些发现， 特别关注了解纤毛基因的遗传变异如何扰乱纤毛运输和 胰岛素受体的下游信号转导。通过仔细使用细胞和分子生物学方法 包括信号分析、RNAseq和活细胞成像，我将量化胰岛素受体信号的差异 以及当目标1中确定的单个纤毛候选基因被敲除时胰岛素受体的纤毛运输 在细胞模型中。拟议工作的完成将增进我们对纤毛生物学和 常见疾病遗传学，为患者风险评估开辟了新的途径，并有可能识别新的 糖尿病治疗的治疗靶点。此外，方法论和范式的应用 这一计划的发展无疑将有助于阐明多种其他常见疾病的发病机制 可归因于孟德尔病基因和纤毛内外的细胞过程。

PROJECT SUMMARY Rare genetic defects of the primary cilium can cause a number of phenotypically-overlapping Mendelian syndromes, known as ciliopathies. Intriguingly, many ciliopathy patients show signs of impaired glucose homeostasis, with accumulating evidence suggesting that insulin receptor must be specifically trafficked into the cilium to initiate insulin-dependent signaling cascades. However, we do not yet know if variants in ciliary genes increase an individual’s risk for diabetes more generally, nor do we understand the mechanisms by which ciliary genes might affect insulin receptor signaling to cause pathology. The goal of this proposal is to fill this gap in our knowledge using complementary bioinformatic and cell biologic approaches. My clinical and scientific expertise in ciliary biology and human genetics, coupled with my exciting preliminary results, rigorous research plan, and outstanding mentorship team, provide a solid foundation for this project. In completing the proposed work, specifically with its focus on providing me additional training in the field of bioinformatics and in lab management, I will be perfectly poised to begin my independent academic career as a physician scientist seeking to better understand the genetics and biology of ciliary dysfunction in human disease. I have secured the complete support of my institution, and will benefit greatly from the unparalleled resources and mentorship available at both the University of Pennsylvania and the Children’s Hospital of Philadelphia over the course of this award. I hypothesize that variants in ciliary genes increase risk for diabetes by perturbing cell signaling pathways essential to normal insulin signaling. My preliminary analyses in the UK and Penn Medicine Biobanks have already identified numerous ciliary genes and variants significantly associated with glucose and hemoglobin A1c levels, and demonstrate significant changes in ciliary gene expression in response to insulin. I will further investigate these associations with two complementary sets of analyses. Specifically, I will (1) use bioinformatic approaches in large patient datasets to identify and characterize ciliary genes and pathways that confer increased risk for type 2 diabetes phenotypes and (2) validate these findings in cell models of disease, with a specific focus on understanding how genetic variation in ciliary genes perturbs the ciliary transport and downstream signaling of insulin receptor. Through the careful use of cell and molecular biologic approaches including signaling assays, RNAseq, and live cell imaging, I will quantify differences in insulin receptor signaling and ciliary transport of insulin receptor when individual ciliary candidate genes, identified in Aim 1, are knocked down in cell models. The completion of the proposed work will advance our knowledge of cilium biology and common disease genetics, open new avenues for patient risk assessment, and has the potential to identify novel therapeutic targets for the treatment of diabetes. Additionally, the application of the methodology and paradigm developed in this plan will undoubtedly help with the elucidation of multiple other common disease mechanisms attributable to Mendelian disease genes and cellular processes both within and outside of the cilium.