Biochemical and cell biological mechanisms of signal transduction through the Hedgehog pathway

Hedgehog 通路信号转导的生化和细胞生物学机制

• 批准号: 9980196
• 负责人: RAJAT ROHATGI
• 金额: $ 67.44万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9980196
• 关键词: Area; Binding Sites; Biochemical; Biological; Cell Communication; Cell Nucleus; Cells; Chemicals; Cilia; Clinical; Collaborations; Congenital Abnormality; Degenerative Disorder; Development; Disease; Embryology; Erinaceidae; Event; Family; Gene Activation; Genes; Glioblastoma; Human Development; Inherited; Integral Membrane Protein; Investigation; Ligand Binding; Ligands; Lipids; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Membrane; Membrane Proteins; Monitor; Mutation; Natural regeneration; Organelles; Pathway interactions; Pharmaceutical Preparations; Phenotype; Phosphorylation; Play; Process; Proteins; Proteomics; Regenerative Medicine; Regulation; Research; Research Personnel; Role; SMO gene; Signal Transduction; Structure; Surface; Synthesis Chemistry; System; Transcription Coactivator; Transcriptional Activation; Vertebrates; Work; ciliopathy; comparative; developmental disease; human disease; human tissue; programs; protein complex; protein transport; public health relevance; receptor; repaired; response; small molecule; smoothened signaling pathway; structural biology; tissue regeneration; tool; transcription factor

 DESCRIPTION (provided by applicant): The Hedgehog (Hh) pathway is a cell-cell communication system that plays important roles in development, regeneration and cancer. Hh signaling is orchestrated in vertebrates at primary cilia, antenna-like organelles that project fro the surfaces of most cells in our bodies and serve as signaling centers in development. Mutations in cilia genes cause a number of inherited human diseases called "ciliopathies," many of which are characterized by birth defects and other phenotypes attributable to aberrant Hh signaling. Despite its importance as a target in cancer and regenerative medicine, many of the steps in Hh signaling remain poorly understood at the biochemical and cell biological level. My research program is focused on the major unsolved mechanistic questions in the vertebrate Hh pathway, with a particular emphasis on ciliary mechanisms that mediate signal propagation and transcriptional activation. The Hh signal is transmitted across the membrane by the 7-pass transmembrane protein Smo, the target for all anti-Hh drugs in clinical use. However, we do not understand the mechanism by which Smo is activated in response to Hh ligands, nor do we know how activated Smo in turn signals to the Glioblastoma (Gli) family of transcription factors. Primary cilia play an important role in both steps- Hh ligands promote the accumulation of Smo in the ciliary membrane, a critical step in signaling that eventually leads to the activation of Gl proteins as they traffic through the ciliary compartment. Delineating this mechanism will provide a valuable paradigm for how TM receptors relay signals from the ciliary membrane to the nucleus. Three major questions under investigation are (1) how Smo is activated by the main Hh receptor Patched 1 through endogenous small molecule ligands, (2) how activated Smo in the ciliary membrane transmits signals to the Gli proteins and (3) how Gli proteins are converted into transcriptional activators. We have made progress in each of these areas. Using new chemical tools to probe the interaction between Smo and oxysterols, endogenous lipids that can activate Hh signaling, we identified and structurally characterized a previously unknown ligand-binding site with regulatory potential in Smo. Using comparative proteomics, we identified a ciliary membrane protein complex that engages Smo at cilia in response to Hh signals and is required for Smo signaling. Finally, we have characterized both dynamic phosphorylation and protein association events that play critical role in Gli activation. Our work is supported by productive collaborations with investigators who have expertise in synthetic chemistry, structural biology, mass spectrometry and embryology. The successful completion of this project will provide (1) an answer to the question of Smo regulation, perhaps the longest-standing mystery in the Hh pathway, (2) an understanding of how ciliary protein trafficking drives signaling and how these processes are corrupted in ciliopathies, and (3) new strategies to monitor and modulate the pathway in Hh-related diseases.

 描述（由申请人提供）：Hedgehog（Hh）通路是一种细胞间通讯系统，在发育、再生和癌症中发挥重要作用。在脊椎动物中，Hh信号是在初级纤毛中协调的，初级纤毛是一种类似触角的细胞器，从我们身体中大多数细胞的表面伸出，并在发育中作为信号中心。纤毛基因的突变导致许多遗传性人类疾病，称为“纤毛病”，其中许多的特征是出生缺陷和其他表型归因于异常的Hh信号传导。尽管它作为癌症和再生医学中的靶点很重要，但Hh信号传导中的许多步骤在生物化学和细胞生物学水平上仍然知之甚少。我的研究项目主要集中在脊椎动物Hh通路中未解决的主要机制问题，特别强调介导信号传播和转录激活的纤毛机制。Hh信号通过7次跨膜蛋白Smo跨膜传递，Smo是临床使用的所有抗Hh药物的靶标。然而，我们不了解Smo响应于Hh配体而被激活的机制，我们也不知道激活的Smo如何反过来向胶质母细胞瘤（Gli）家族的转录因子发出信号。初级纤毛在这两个步骤中起重要作用- Hh配体促进Smo在睫状体膜中的积累，这是信号传导中的关键步骤，当它们通过睫状体隔室运输时，其最终导致GI蛋白的活化。阐明这一机制将为TM受体如何将信号从睫状体膜传递到细胞核提供一个有价值的范例。研究中的三个主要问题是（1）Smo如何通过内源性小分子配体被主要Hh受体Patched 1激活，（2）睫状膜中激活的Smo如何将信号传递给Gli蛋白，以及（3）Gli蛋白如何转化为转录激活因子。我们在这些领域都取得了进展。使用新的化学工具来探测Smo和氧化固醇，内源性脂质，可以激活Hh信号之间的相互作用，我们确定和结构特征的一个以前未知的配体结合位点与Smo的监管潜力。使用比较蛋白质组学，我们确定了一个睫状膜蛋白复合物，从事Smo在纤毛响应Hh信号，并需要Smo信号。最后，我们的特点是动态磷酸化和蛋白质协会的事件，发挥关键作用Gli激活。我们的工作得到了与具有合成化学、结构生物学、质谱和胚胎学专业知识的研究人员的富有成效的合作的支持。该项目的成功完成将提供（1）Smo调节问题的答案，这可能是Hh通路中存在时间最长的谜团，（2）了解纤毛蛋白运输如何驱动信号传导以及这些过程如何在纤毛病中被破坏，以及（3）监测和调节Hh相关疾病通路的新策略。