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Molecular mechanisms of Hedgehog receptor function

Hedgehog受体功能的分子机制

基本信息

批准号：
10737476
负责人：
PHILIP A BEACHY
金额：
$ 40.45万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-08-15 至 2027-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10737476
关键词：
Acceleration Address Adult Affinity Anatomy Basal cell carcinoma Binding Binding Proteins Biochemical Biological Biological Assay Bladder Blood - brain barrier anatomy Cations Cause of Death Cell membrane Cells Cholesterol Chronic Chronic Obstructive Pulmonary Disease Cilia Complex Cryoelectron Microscopy Disease Down Syndrome Embryonic Pattern Specification Energy-Generating Resources Erinaceidae Event Family Family member Funding G-Protein-Coupled Receptors Hydrophobicity In Vitro Inflammatory Bowel Diseases Injury Intestines Ligands Link Lipids Lung Maintenance Malignant Neoplasms Measures Mediating Membrane Metals Molecular Molecular Conformation Muscle Natural regeneration Organ Pancreas Pathology Pathway interactions Pattern Pharmaceutical Preparations Play Porifera Postembryonic Primary Neoplasm Proliferating Protein Inhibition Proteins Regulation Resolution Role SHH gene Signal Induction Signal Pathway Signal Transduction Signaling Protein Site Specific qualifier value Sterols Structure Surface Therapeutic Tissue Differentiation Tissues Transducers Work antagonist bone cancer therapy cell type embryo tissue hedgehog signal transduction in vivo inhibitor member morphogens nanobodies neoplastic cell novel novel strategies polypeptide prevent protein complex protein protein interaction protein structure receptor receptor function regenerative repaired resistance mutation smoothened signaling pathway stem cells tissue stem cells transmission process tumor tumor growth tumor progression

项目摘要

Project Summary Hedgehog (Hh) signaling specifies the embryonic tissue pattern of many metazoan organs and maintains this tissue pattern post-embryonically by regulating the expression of proliferation- or differentiation-inducing signals that target adult tissue stem or progenitor cells. Drugs developed to block Hh pathway activity, based on our previous work, have received FDA approval for treatment of ectodermally-derived cancers, such as basal cell carcinoma. In pancreatic, bladder, and other cancers of endodermal origin, Hh pathway activity in tumor- associated stroma presents a barrier to tumor growth and progression, thus suggesting pathway activation rather than inhibition as a therapeutic approach. In addition, pathway activation has a beneficial regenerative role in bone and muscle repair, in reducing pathology associated with inflammatory bowel disease, and in preventing or ameliorating injury and breach of the blood-brain-barrier, among other emerging biological activities. On the other hand, chronic low-level elevation of pathway activity in the lung, as is associated with reduced expression of the Hh pathway inhibitor Hhip (Hh-interacting protein), is genetically linked to chronic obstructive pulmonary disease (COPD), the third leading cause of death worldwide. During the previous funding period for this project we utilized protein structure determination and biochemical and cell biological approaches to establish the molecular mechanism of Hh signaling, in which Hh binding to its receptor Patched1 (Ptch1) activates the pathway by alleviating Ptch1-mediated suppression of the essential transducer and GPCR family member, Smoothened (Smo). We found that cholesterol is the crucial link between Ptch1 and Smo, that cholesterol in the inner leaflet of the membrane is decreased by Ptch1 transport activity, and that Hh binding to Ptch1 blocks this transport activity. These events critically regulate pathway activity, as conformational switching of Smo to its active state requires entry and binding of a sterol from the inner leaflet of the membrane into a central cavity within the Smoothened seven-transmembrane bundle. We also showed how the Dispatched1 (Disp1) transporter, structurally related to Ptch1, uses Na+ flux to power its export and packaging of the dually lipid modified Sonic hedgehog protein signal (ShhNp), enabling it to move through tissues as a soluble morphogen in complex with its carrier Scube2. We propose here to deepen our understanding of Hh signal transduction and pathway regulation by establishing the energy sources and the step-by-step lipid- handling mechanisms of the Ptch1 and Disp1 transporters. We will determine the structure of the ShhNp:Scube2 morphogen, and the mechanism of its release from Disp1. Finally we plan to elucidate the mechanism of Hh signal antagonism by Hhip, using cryo-EM to determine the high-resolution structure and functionally dissect a membrane-associated tent-like Hhip multimeric complex that occludes all receptor-interacting surfaces of the Hh protein. Our findings may provide a basis for new approaches to therapeutic modulation of Hh pathway activity.

项目概要 Hedgehog (Hh) 信号传导指定许多后生动物器官的胚胎组织模式并维持这种模式通过调节增殖或分化诱导信号的表达来调节胚胎后组织模式靶向成体组织干细胞或祖细胞。根据我们的研究，开发出阻断 Hh 通路活性的药物之前的工作已获得 FDA 批准用于治疗外胚层衍生癌症，例如基底细胞癌癌。在胰腺癌、膀胱癌和其他内胚层来源的癌症中，肿瘤中的 Hh 通路活性相关基质对肿瘤生长和进展构成障碍，因此表明通路激活而不是比抑制作为治疗方法。此外，通路激活具有有益的再生作用骨骼和肌肉修复，减少与炎症性肠病相关的病理，并预防或改善损伤和血脑屏障的破坏，以及其他新兴的生物活性。上另一方面，肺部通路活性的慢性低水平升高，与表达减少相关 Hh 通路抑制剂 Hhip（Hh 相互作用蛋白）的基因与慢性阻塞性肺疾病有关慢性阻塞性肺病 (COPD)，是全球第三大死因。在该项目的上一个资助期间，我们利用了蛋白质结构测定和生化和细胞生物学方法来建立 Hh 信号传导的分子机制，其中 Hh 与其结合受体 Patched1 (Ptch1) 通过减轻 Ptch1 介导的基本抑制来激活该通路传感器和 GPCR 家族成员 Smoothened (Smo)。我们发现胆固醇是两者之间的重要联系 Ptch1 和 Smo，Ptch1 转运活性降低了膜内小叶中的胆固醇， Hh 与 Ptch1 的结合会阻断这种转运活动。这些事件严格调节通路活性，如 Smo 构象转变为其活性状态需要从内部小叶进入并结合甾醇将膜放入平滑的七跨膜束内的中央腔中。我们还展示了如何 Dispatched1 (Disp1) 运输机在结构上与 Ptch1 相关，使用 Na+ 通量为其输出和包装提供动力双重脂质修饰的音刺猬蛋白信号 (ShhNp)，使其能够作为可溶性形态发生素与其载体 Scube2 形成复合物。我们在这里建议加深对Hh的理解通过建立能量来源和逐步脂质-信号转导和途径调节 Ptch1 和 Disp1 转运蛋白的处理机制。我们将确定ShhNp:Scube2的结构形态发生素及其从 Disp1 中释放的机制。最后我们计划阐明Hh的机制 Hhip 的信号拮抗作用，使用冷冻电镜确定高分辨率结构并从功能上剖析膜相关的帐篷状 Hhip 多聚体复合物，封闭 Hh 的所有受体相互作用表面蛋白质。我们的研究结果可能为 Hh 通路活性的治疗调节新方法提供基础。

项目成果

期刊论文数量（10）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Itraconazole inhibits enterovirus replication by targeting the oxysterol-binding protein.

DOI：
10.1016/j.celrep.2014.12.054
发表时间：
2015-02-03
期刊：
Cell reports
影响因子：
8.8
作者：
Strating JR;van der Linden L;Albulescu L;Bigay J;Arita M;Delang L;Leyssen P;van der Schaar HM;Lanke KH;Thibaut HJ;Ulferts R;Drin G;Schlinck N;Wubbolts RW;Sever N;Head SA;Liu JO;Beachy PA;De Matteis MA;Shair MD;Olkkonen VM;Neyts J;van Kuppeveld FJ
通讯作者：
van Kuppeveld FJ

Simultaneous measurement of smoothened entry into and exit from the primary cilium.

同时测量初级纤毛的平滑进入和退出。

DOI：
10.1371/journal.pone.0104070
发表时间：
2014
期刊：
PloS one
影响因子：
3.7
作者：
Kim,Jynho;Hsia,ElaineYC;Kim,James;Sever,Navdar;Beachy,PhilipA;Zheng,Xiaoyan
通讯作者：
Zheng,Xiaoyan

Hedgehog pathway modulation by multiple lipid binding sites on the smoothened effector of signal response.

Hedgehog 通路通过信号响应平滑效应器上的多个脂质结合位点进行调节。

DOI：
10.1016/j.devcel.2013.07.015
发表时间：
2013-08-26
期刊：
DEVELOPMENTAL CELL
影响因子：
11.8
作者：
Myers, Benjamin R.;Sever, Navdar;Chong, Yong Chun;Kim, James;Belani, Jitendra D.;Rychnovsky, Scott;Bazan, J. Fernando;Beachy, Philip A.
通讯作者：
Beachy, Philip A.

The role of ciliary trafficking in Hedgehog receptor signaling.