Mechanisms and function of the microtubule podosome connection in osteoclasts

描述（由申请人提供）：破骨细胞吸收增加是多种骨病的原因，无论是全身性（骨质疏松症、佩吉特病、甲状旁腺功能亢进等）还是局部（类风湿关节炎、牙周病、多发性骨髓瘤、肿瘤性骨溶解等）。尽管存在几种抗吸收药物，但不良影响的证据正在出现。因此，更好地了解破骨细胞（OC）和骨吸收的生物学仍然具有最大的临床重要性。我们的实验室已经投入了大量的资金来详细了解骨吸收和骨吸收的分子生物学。这项工作的最显著特征之一，得到了许多其他实验室的支持，是证明了OC与骨的附着至关重要，由一种特殊的粘附结构介导，即含有肌动蛋白的足体，在OC中，它被专门组织在外周带（肌动蛋白环）中，当OC附着在骨上并主动吸收时，最终在密封区中。足体带的形成是oc特有的特征。一项重要的突破是证明了微管（MTs）在将足小体组织成肌动蛋白环和封闭区，从而在骨吸收中所起的关键作用（Destaing等人，2004；jill - henn等人，2007；Purev等人，2009）。本应用程序的主要目的是进一步阐明MTs连接和调节足小体的分子机制，特别是从团簇到外周带的转变，以及骨吸收。我们的初步数据揭示了MTs、+Tip蛋白EB1、接触层和足小体之间的功能相互作用，这是形成足小体带所必需的。我们的建议侧重于MTs +末端的分子调控及其与OCs足质体中肌动蛋白和肌动蛋白调节蛋白的相互作用。这将有助于理解调节足小体的定位和周转及其在外周带中的组织的机制，外周带是oc特异性的，是建立封闭区的必要步骤，本身是骨吸收所必需的。我们建议：1-确定+TIPs蛋白EB1在连接MTs与肌动蛋白动力学中的作用。2-确定接触蛋白（及其密切同源物HS1）及其与EB1的相互作用在足小体MTs和肌动蛋白动力学之间的联系中的作用。3-分析Src和HDAC6的作用，即酪氨酸磷酸化和乙酰化，在MTs和肌动蛋白动力学之间的联系。考虑到足小体和侵足体的相似性，这些研究可能为调节骨吸收开辟新的治疗途径，对骨质疏松症、溶骨性疾病和癌症的治疗有潜在的影响。

DESCRIPTION (provided by applicant): Increased osteoclastic resorption is responsible for multiple bone diseases, whether systemic (osteoporosis, Paget's disease, hyper-parathyroidism for instance) or local (rheumatoid arthritis, periodontal disease, multiple myeloma, tumor osteolysis etc.). Despite the existence of several anti- resorptive drugs, evidence of unwanted effects is emerging. It therefore remains of the utmost clinical importance to better understand the biology of the osteoclast (OC) and of bone resorption. Our laboratory has been heavily invested in the detailed molecular understanding of OC biology and bone resorption. One of the most salient features of this work, supported by many other laboratories, is the demonstration of the critical importance of the attachment of the OC to bone, mediated by a specialized adhesion structure, the actin-containing podosome, which in OCs is specifically organized in a peripheral belt (the actin ring) and ultimately in a sealing zone when OCs are on bone and actively resorbing. The formation of a podosome belt is an OC-specific feature. An important breakthrough has been the demonstration of the critical role played by microtubules (MTs) in the organization of podosomes into an actin ring and a sealing zone and consequently in bone resorption (Destaing et al. 2004; Gil-Henn et al., 2007; Purev et al., 2009). The primary goal of this application is to further elucidate the molecular mechanisms by which MTs are connected to and regulate the podosomes, and in particular the transition from clusters to a peripheral belt, and bone resorption. Our preliminary data revealed a functional interaction between MTs, the +Tip protein EB1, cortactin and podosomes, required for the formation of podosome belts. Our proposal focuses on the molecular regulation of the + ends of MTs and of their interaction with actin and actin-regulating proteins in podosomes in OCs. This will help understand the mechanisms that regulate the localization and turnover of podosomes and their organization in a peripheral belt, an OC-specific and necessary step in the establishment of the sealing zone, itself required for bone resorption. We propose to: 1- Determine the role of the +TIPs protein EB1 in linking MTs to actin dynamics in podosomes. 2- Determine the role of cortactin (and its close homolog HS1) and its interaction with EB1 in the link between MTs and actin dynamics in podosomes. 3- Analyze the role of Src and HDAC6, i.e. Tyrosine phosphorylation and acetylation, in the link between MTs and actin dynamics in podosomes. The studies may open new therapeutic avenues to regulate bone resorption, with potential impact on the treatment of osteoporosis, osteolytic diseases and also cancer, given the similarities between podosomes and invadopodia. PUBLIC HEALTH RELEVANCE: Increased osteoclastic resorption is responsible for multiple bone diseases, whether systemic (osteoporosis, Paget's disease, hyper-parathyroidism for instance) or local (rheumatoid arthritis, periodontal disease, multiple myeloma, primary or metastatic tumor osteolysis etc.). The primary goal of this application is to further elucidate the molecular mechanisms by which osteoclast attach to and resorb bone to better understand the biology of the osteoclast and of bone resorption to design new and safer drugs.