Biochemical Mechanisms of Hedgehog Signaling

Hedgehog 信号转导的生化机制

• 批准号: 7917083
• 负责人: RAJAT ROHATGI
• 金额: $ 24.9万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2012-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7917083
• 关键词: Actins; Adult; Affect; Affinity Chromatography; Antibodies; Binding; Biochemical; Biological; Biological Assay; Brain; Cancer Patient; Cancer cell line; Caring; Cell Surface Extensions; Cell division; Cell physiology; Cell surface; Cells; Cilia; Congenital Abnormality; Cytoskeleton; Defect; Detection; Development; Doctor of Philosophy; Educational process of instructing; Embryo; Erinaceidae; Event; Fibroblasts; Genetic Epistasis; Genetic Transcription; Goals; Hereditary Malignant Neoplasm; Homeostasis; Human; Image; Laboratories; Lead; Link; Location; Lung; Maintenance; Malignant Neoplasms; Mammals; Mass Spectrum Analysis; Mentors; Methods; Microscopy; Modeling; Molecular; Morphogenesis; Mus; Mutation; Optics; Organ; Pancreas; Pathogenesis; Pathway interactions; Phase; Phosphorylation; Play; Positioning Attribute; Post-Translational Protein Processing; Prostate; Protein Biochemistry; Protein Dynamics; Proteins; Regulation; Research; Role; Sensory; Signal Pathway; Signal Transduction; Skin; Stem cells; Structure; Surface; Syndrome; Techniques; Time; Tissue Expansion; Tissues; Training; Upper digestive tract structure; Work; base; body system; cancer cell; carcinogenesis; experience; hedgehog signal transduction; human SMO protein; human disease; neoplastic cell; novel; novel therapeutics; oncology; protein complex; receptor; smoothened signaling pathway; tissue culture; tool; tumorigenesis

DESCRIPTION (provided by applicant): Aberrant activation of the Hedgehog signaling pathway has been implicated in the genesis and maintenance of human cancers that develop in multiple organ systems, including the skin, brain, lung, prostate, and pancreas. It is hypothesized that the pathway drives tumorigenesis by causing the activation and expansion of tissue stem cells. There are significant gaps in our understanding of how the mammalian Hedgehog signal is received and transduced. Deciphering the detailed biochemical mechanism of Hedgehog signaling will allow the development of novel therapeutics and preventative strategies for these lethal cancers. During my PhD training with Dr. Marc Kirschner, I used a combination of protein biochemistry and microscopy to dissect a pathway that links cell surface signals to the actin cytoskeleton. To apply a similar biochemical and cell-biological approach to Hedgehog signaling, I have developed novel antibodies and confocal microscopy- based assays to study the dynamics and interactions of proteins in the pathway. Based on this work, I have constructed a new model for Hedgehog signaling that highlights the importance of the primary cilium, a tiny projection found on the surface of most cells that has been recently implicated in the Hedgehog pathway and in human disease. In the independent phase of this proposal, I plan to use the above tools to understand how localization of the receptor, Patched 1, to the primary cilium affects its ability to sense the Sonic Hedgehog signal and to activate downstream signaling in mouse embryonic fibroblasts and human tumor cells. I will also use an unbiased immunoaffinity purification approach to discover novel interacting proteins and post-translational modifications that link Hedgehog pathway components to the primary cilium. In the K99 phase of this proposal, Dr. Matthew Scott, a pioneer in the analysis of Hedgehog signaling in human and mouse cancer, will serve as mentor. My time in his laboratory will provide a critical opportunity to gain experience in techniques for the analysis of Hedgehog signal transduction in tissue culture fibroblasts, tumor cells and mice and to develop optical and biochemical probes for the proposed imaging and protein- interaction analysis. Most importantly, it will put me in an ideal position for a tenure-track position in an oncology department, where I plan to spend ~80-90% of my time in research and teaching and 10-20% of my time in the care of cancer patients.

描述（由申请人提供）：Hedgehog 信号通路的异常激活与人类癌症的发生和维持有关，这些癌症在多个器官系统中发展，包括皮肤、大脑、肺、前列腺和胰腺。据推测，该途径通过引起组织干细胞的激活和扩增来驱动肿瘤发生。我们对哺乳动物刺猬信号如何接收和转导的理解存在重大差距。破译 Hedgehog 信号传导的详细生化机制将有助于开发针对这些致命癌症的新疗法和预防策略。在 Marc Kirschner 博士的指导下进行博士培训期间，我结合使用蛋白质生物化学和显微镜来剖析将细胞表面信号与肌动蛋白细胞骨架连接起来的途径。为了将类似的生化和细胞生物学方法应用于 Hedgehog 信号传导，我开发了新型抗体和基于共聚焦显微镜的测定法来研究该通路中蛋白质的动态和相互作用。基于这项工作，我构建了一个新的 Hedgehog 信号传导模型，该模型强调了初级纤毛的重要性，初级纤毛是大多数细胞表面发现的微小突起，最近与 Hedgehog 通路和人类疾病有关。在该提案的独立阶段，我计划使用上述工具来了解受体（Patched 1）在初级纤毛上的定位如何影响其感知 Sonic Hedgehog 信号以及激活小鼠胚胎成纤维细胞和人类下游信号传导的能力肿瘤细胞。我还将使用无偏免疫亲和纯化方法来发现将 Hedgehog 通路成分与初级纤毛连接起来的新型相互作用蛋白和翻译后修饰。在该提案的K99阶段，人类和小鼠癌症中Hedgehog信号传导分析的先驱Matthew Scott博士将担任导师。我在他的实验室的时间将为我提供一个重要的机会，让我获得组织培养成纤维细胞、肿瘤细胞和小鼠中 Hedgehog 信号转导分析技术的经验，并为所提出的成像和蛋白质相互作用分析开发光学和生化探针。最重要的是，这将使我处于肿瘤科终身职位的理想位置，我计划将大约 80-90% 的时间用于研究和教学，10-20% 的时间用于护理的癌症患者。