Biochemical mechanisms of Hedgehog signal transduction through primary cilia

Hedgehog通过初级纤毛信号转导的生化机制

基本信息

批准号：
10447446
负责人：
David R Raleigh
金额：
$ 20.19万
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：
2022
资助国家：
美国
起止时间：
2022-07-01 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10447446
关键词：
Address Adult Animals Antibodies Applications Grants Architecture Ascorbic Acid Binding Biochemical Biological Assay Biophysical Process CRISPR interference CRISPR/Cas technology Cell Proliferation Cells Cilia Congenital Disorders Critical Pathways Data Development Disease Erinaceidae Event Family Future Gene Expression Genes Genetic Genetic Transcription Goals Health Homeostasis Human Human Biology Immunofluorescence Immunologic Investigation Kinetics LLC-PK1 Cells Label Ligands Light Lipid Binding Lipids Luciferases Malignant Neoplasms Mass Spectrum Analysis Measures Mediating Membrane Transport Proteins Molds Molecular Normal Cell Oncogenic Output Pathway interactions Patients Peroxidases Phenotype Physiological Proteins Proteomics Reaction Regulation Reporter Research Project Grants Role SHH gene Sampling Signal Transduction Signaling Molecule Specific qualifier value Surface Testing Tissues Tumor Cell Line Vertebrates cell type experimental study functional genomics hedgehog signal transduction high dimensionality improved in vivo innovation lipidomics migration neoplastic cell programs response smoothened signaling pathway spatiotemporal trafficking transcription factor

项目摘要

Project summary Spatiotemporal regulation of signal molecules is critical for development and adult tissue homeostasis. Indeed, misactivation or mislocalization of signaling events can cause congenital disorders and cancers, and understanding how information transfer is regulated in cells is essential for understanding and improving human health. The Hedgehog pathway is conserved across metazoan animals where it controls cell proliferation, differentiation, migration, and homeostasis. In vertebrates, Hedgehog signals are transduced through primary cilia that project from the surface of most cells, including cells in cancers driven by misactivation of the Hedgehog pathway. Cilia are required for vertebrate Hedgehog signaling for reasons that remain unknown. Our central hypothesis is that the ciliary microenvironment enables protein and lipid interactions that are necessary for Hedgehog signal transduction. Our lab has identified an important role for cilia-associated lipids that bind to Smoothened, accumulate Smoothened to cilia, and activate the Hedgehog pathway. When the Hedgehog pathway is off, Smoothened accumulation and activity in cilia is inhibited by Patched1. How Patched1 and Smoothened accumulate and function in cilia, and how Patched1 inhibits Smoothened, are poorly understood despite increasing recognition that these activities of the Hedgehog pathway are critical in health and disease. To address these key gaps in our understanding of human biology, the objectives of this proposal are to define the protein interactions that are necessary for Patched1 and Smoothened accumulation and activity in primary cilia, and to determine if Patched1 inhibited Smoothened through compartmentalization of ciliary lipids. Recently, we have begun to appreciate that the Hedgehog pathway can gather information directly from the ciliary microenvironment. Our discovery of ciliary lipids that specify Hedgehog pathway output establishes a new paradigm for information input onto the Hedgehog transcriptional program, one in which changes in the ciliary microenvironment reprogram the Hedgehog response. In this proposal, we will systematically examine the dynamic repertoire of protein and lipid interactions in cilia that specify the Hedgehog pathway across normal and tumor cells. We will leverage recent technical advances in proteomic proximity-labeling mass spectrometry, lipidomic mass spectrometry, and functional genomics to interpret high-dimensional landscapes of cell states induced by Hedgehog pathway activation. The long-term goal of this proposal is to understanding mechanisms underlying Hedgehog signal transduction well enough to develop new treatments for Hedgehog-associated cancers. To do so, the data generated by this proposal will establish a rationale to interrogate these mechanisms in vivo and in samples from human patients with Hedgehog-associated cancers. More broadly, this proposal will elucidate why cilia are required for Hedgehog signaling and how information transfer is regulated in cells, an elusive and fundamental question of human biology.

项目概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。事实上，信号事件的错误激活或错误定位可能导致先天性疾病和癌症，并且了解细胞中信息传递的调节方式对于理解和改善人类的行为至关重要健康。 Hedgehog 通路在后生动物中是保守的，它控制细胞增殖，分化、迁移和稳态。在脊椎动物中，Hedgehog 信号通过初级信号转导从大多数细胞表面突出的纤毛，包括由刺猬失活驱动的癌症细胞途径。脊椎动物 Hedgehog 信号传导需要纤毛，其原因尚不清楚。我们的中央假设是纤毛微环境能够实现蛋白质和脂质的相互作用，这是维持细胞生长所必需的。刺猬信号转导。我们的实验室已经确定了纤毛相关脂质的重要作用，这些脂质与 Smoothened，积累Smoothened至纤毛，激活Hedgehog通路。当刺猬途径关闭，纤毛中平滑的积累和活动受到 Patched1 的抑制。如何修补1和人们对纤毛中 Smoothened 的积累和功能以及 Patched1 如何抑制 Smoothened 知之甚少尽管人们越来越认识到 Hedgehog 通路的这些活动对于健康和疾病至关重要。为了解决我们对人类生物学理解中的这些关键差距，该提案的目标是定义 Patched1 和 Smoothened 积累和活性所需的蛋白质相互作用初级纤毛，并确定 Patched1 是否通过纤毛脂质的区室化抑制 Smoothened。最近，我们开始意识到 Hedgehog 通路可以直接从睫状体微环境。我们对指定 Hedgehog 通路输出的纤毛脂质的发现建立了一个新的 Hedgehog 转录程序的信息输入范式，其中睫状体的变化微环境重新编程刺猬反应。在本提案中，我们将系统地审查纤毛中蛋白质和脂质相互作用的动态库，指定了跨正常和肿瘤细胞。我们将利用蛋白质组邻近标记质谱的最新技术进展，脂质组学质谱和功能基因组学来解释细胞状态的高维景观由 Hedgehog 通路激活诱导。该提案的长期目标是了解 Hedgehog 信号的潜在机制转导效果足以开发针对刺猬相关癌症的新疗法。为此，数据该提案产生的结果将建立在体内和样本中质疑这些机制的基本原理患有刺猬相关癌症的人类患者。更广泛地说，该提案将阐明为什么纤毛是 Hedgehog 信号传导以及细胞中信息传递的调节方式所需，这是一个难以捉摸且基本的问题人类生物学问题。