Regulation of Signaling in Osteoclast Bone Resorption

描述（由申请人提供）：本提案的长期目标是阐明参与肌动蛋白环形成和骨吸收的分子机制。骨吸收是骨重塑的第一步。肌动蛋白环的形成已被证明是破骨细胞有效骨吸收的先决条件。我们实验室最近的初步研究表明，N-WASP-Arp2/3复合体可能在破骨细胞肌动蛋白环的形成中起作用。酪氨酸激酶如PYK2和Src参与了N-WASP和N-WASP相关磷酸化蛋白的磷酸化。此外，酪氨酸磷酸酶（PTP-PEST）在调节N-WASP及其相关信号分子的酪氨酸磷酸化中具有独特的作用。因此，我们假设：1。由Cdc42、PtdIns P2 （PIP2）和激酶(s)协同激活的N-WASP可以刺激Arp2/3介导的破骨细胞肌动蛋白聚合和肌动蛋白环的形成。2. Src/PTP-PEST调控N-WASP和相关信号蛋白的酪氨酸磷酸化是肌动蛋白环和骨吸收中肌动蛋白重塑所必需的。因此，我们的具体目标是：1。确定参与N-WASP激活和肌动蛋白环形成的信号转导机制。2. 通过PTP-PEST测定酪氨酸磷酸化，肌动蛋白环形成和骨吸收的调节。本修订更新申请的目的是确定肌动蛋白环形成的潜在分子机制。为了在细胞和分子水平上促进对骨吸收机制的理解，将使用不同的方法。HIV-TAT或腺病毒介导的N-WASP、PTP-PEST和激酶（Src和PYK2）进入破骨细胞，以确定参与N-WASP- arp2 /3复合物和肌动蛋白环形成的信号转导机制。PTP-PEST与N-WASP的结合位点将通过传递来自PTP-PEST和N-WASP富含脯氨酸区域的tat融合寡肽来表征。我们将分析上述治疗对肌动蛋白环形成和骨吸收的影响。鉴定阻碍破骨细胞功能的肽将有助于开发针对骨质疏松症、牙周病和骨关节炎等疾病的破骨细胞肌动蛋白环形成和骨吸收的药理学药物。

DESCRIPTION (provided by applicant): The long-term objective of this proposal is to elucidate the molecular mechanisms involved in actin ring formation and bone resorption. Bone resorption is the first step in bone remodeling. Actin ring formation has been shown to be a prerequisite for efficient bone resorption in osteoclasts. Recent preliminary studies from our laboratory suggest that N-WASP-Arp2/3 complex may have a role in the osteoclast actin ring formation. Tyrosine kinases such as PYK2 and Src are involved in the phosphorylation of N-WASP and N-WASP associated phosphoproteins. Moreover, tyrosine phosphatase (PTP-PEST) has a unique role in the modulation of tyrosine phosphorylation of N-WASP and the associated signaling molecules. We therefore hypothesize: 1. N-WASP coordinately activated by Cdc42, PtdIns P2 (PIP2), and kinase(s) can stimulate Arp2/3 mediated actin polymerization and actin ring formation in osteoclasts. 2. Src/PTP-PEST regulation of tyrosine phosphorylation of N-WASP and the associated signaling proteins is required for actin remodeling in the actin ring and bone resorption. Thus, our Specific Aims are to: 1. Determine the signal transduction mechanisms involved in N-WASP activation and actin ring formation. 2. Determine the regulation of tyrosine phosphorylation, actin ring formation, and bone resorption by PTP-PEST. The goal of this revised renewal application is to identify the underlying molecular mechanisms in actin ring formation. To advance the understanding of the mechanisms of bone resorption at the cellular and molecular level, different approaches will be used. HIV-TAT or adenoviral-mediated delivery of N-WASP, PTP-PEST, and kinases (Src and PYK2) into osteoclasts will be performed to identify the signal transduction mechanisms involved in the formation of N-WASP-Arp2/3 complex and actin ring. The binding sites of PTP-PEST with N-WASP will be characterized by the delivery of TAT-fused oligopeptides derived from proline-rich regions of PTP-PEST and N-WASP. We will analyze the effects of the above-mentioned treatments on actin ring formation and bone resorption. Identification of peptides that impede osteoclast function will be useful in the development of pharmacological agents, targeting osteoclast actin ring formation and bone resorption in disorders such as osteoporosis, periodontal disease, and osteoarthritis.