Understanding Patched1 protein and lipid interactions in cilia

了解纤毛中的 Patched1 蛋白和脂质相互作用

基本信息

批准号：
10315798
负责人：
Vikas daggubati
金额：
$ 3.82万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-01 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10315798
关键词：
Address Adult Affinity Chromatography Animals Antibodies Automobile Driving Basal cell carcinoma Binding Biochemical Biological Assay Biological Models CRISPR interference Cancer cell line Cell Line Cell Maintenance Cell Proliferation Cells Childhood Brain Neoplasm Cilia Co-Immunoprecipitations Congenital Disorders Data Development Electron Microscopy Erinaceidae Family Gene Expression Genes Genetic Genetic Transcription Goals Homeostasis Human Biology Immunofluorescence Immunologic Knowledge LLC-PK1 Cells Label Ligand Binding Light Lipid Binding Lipids Luciferases Malignant Neoplasms Malignant neoplasm of brain Mass Spectrum Analysis Mediating Membrane Transport Proteins Modeling Mutate Pathway interactions Patients Pediatric Neoplasm Peroxidases Proteins Proteomics Recurrence Regulation Reporter Signaling Molecule Sterols Structure Sum Surface System Testing Time Tissues Tumor Suppressor Proteins United States Vertebrates Work ascorbate base cryogenics developmental disease experimental study functional genomics hedgehog signal transduction human disease improved innovation insight lipidomics medulloblastoma medulloblastoma cell line migration neoplastic cell novel novel therapeutics prevent programs smoothened signaling pathway spatiotemporal stem cells tool transcription factor

项目摘要

PROJECT SUMMARY/ABSTRACT Spatiotemporal regulation of signaling molecules is critical for development and adult tissue homeostasis. The Hedgehog pathway, which is conserved across metazoan animals, controls cell proliferation, differentiation, migration, and stem cell maintenance. In vertebrates, Hedgehog signals are transduced through primary cilia that project from the surface of most cells, including cells in cancers that are driven by misactivation of the Hedgehog pathway. The tumor suppressor Patched1 localizes to cilia and inhibits Smoothened, a Hedgehog pathway activator. Upon pathway activation, Patched1 leaves cilia and allows Smoothened to activate the downstream Hedgehog transcriptional program. How Patched1 localizes to cilia and inhibits Smoothened remain unknown. The central hypothesis of this proposal is that a dynamic network of protein interactions allow Patched1 to accumulate in cilia and regulate the ciliary lipid microenvironment to inhibit Hedgehog signal transduction. To test this, the objective of this proposal is to define the ciliary protein interactions necessary for localization of Patched1 to the cilia, and to determine if Patched1 regulates Smoothened by regulating the ciliary lipid microenvironment. To address the gaps in our understanding of Hedgehog signaling, I will leverage recent technical advances in proteomic proximity-labeling mass spectrometry, lipidomic mass spectrometry, and functional genomics using novel model systems I have generated for this proposal. In Aim 1, I will interrogate 5 protein interactors of Patched1 that are associated with human disease and may underlie Patched1 accumulation and activity in primary cilia. In Aim 2, I will define the impact of Patched1 on the ciliary lipid microenvironment. Combined, these aims will elucidate the biochemical mechanism by which Patched1 regulates Hedgehog signaling. Hedgehog pathway misactivation drives medulloblastoma, the most common pediatric brain tumor, and basal cell carcinoma, the most common cancer in the United States. Thus, this proposal will incorporate Hh-associated cancer cell lines to determine if ciliary proteins and lipids underlying Patched1 functions are conserved across developmental and disease contexts. In sum, understanding how Patched1, the most recurrently mutated gene in Hh-associated cancers, inhibits Hedgehog signaling will provide significant insights into human biology and potentially provide avenues for novel therapies.

项目摘要/摘要信号分子的时空调节对于发育和成人组织稳态至关重要。这刺猬途径是跨后生动物保守的，控制细胞增殖，分化，迁移和干细胞维护。在脊椎动物中，刺猬信号通过原发性纤毛转导该项目来自大多数细胞的表面，包括疾病中的细胞。刺猬途径。肿瘤抑制剂修补1位于纤毛并抑制平滑，刺猬途径激活器。途径激活后，Patched1离开纤毛并使平滑以激活下游刺猬转录程序。修补1如何定位到纤毛和抑制平滑的状态未知。该提议的中心假设是蛋白质相互作用的动态网络允许修补1 在纤毛中积聚并调节睫状脂质微环境以抑制刺猬信号转导。到测试这一点，该提案的目的是定义定位所必需的睫状蛋白相互作用对纤毛的修补1修补，并确定修补1是否通过调节纤毛脂质来调节平滑微环境。为了解决我们对刺猬信号的理解的差距，我将利用最近的技术进步在蛋白质组学近端标记的质谱法中，脂质质量质谱法和使用功能基因组学我为此提案生成的新型模型系统。在AIM 1中，我将询问5个蛋白质相互作用者与人类疾病相关的修补1，可能是补丁1的基础1 原发性纤毛。在AIM 2中，我将定义Patched1对睫状脂质微环境的影响。结合了这些 AIMS将通过修补1调节刺猬信号的生化机制来阐明生化机制。刺猬途径误导驱动髓母细胞瘤，最常见的小儿脑肿瘤和基底细胞癌，美国最常见的癌症。因此，该建议将结合HH相关癌细胞系以确定睫状蛋白和脂质是否在跨层次修补1功能的范围发育和疾病环境。总而言之，理解如何修补1，最经常突变的基因在与HH相关的癌症中，抑制刺猬信号将为人类生物学和有可能为新疗法提供途径。