The Role of PLEKHM1 in Osteoclast Function

PLEKHM1 在破骨细胞功能中的作用

• 批准号: 7793419
• 负责人: HAIBO ZHAO
• 金额: $ 7.18万
• 依托单位: UNIV OF ARKANSAS FOR MED SCIS
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-04-01 至 2012-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7793419
• 关键词: Aging; Albers-Schonberg disease; American; Binding; Bone Resorption; Cells; Chloride Channels; Defect; Equilibrium; Gene Targeting; Genes; Genetic; Gray unit of radiation dose; Homeostasis; Human; Incisor; Integrins; Modeling; Mutate; Mutation; Osteoblasts; Osteoclasts; Osteogenesis; Osteoporosis; Patients; Pharmaceutical Preparations; Population; Positioning Attribute; Proteins; Public Health; RNA Interference; Rattus; Resistance; Rodent; Role; SRC gene; Structure; System; TRAF6 gene; Tissues; bone; bone loss; bone mass; bone resorbing activity; cathepsin K; cost; knockout gene; mutant; novel; prevent; small hairpin RNA; vacuolar H+-ATPase

DESCRIPTION (provided by applicant): A tight balance between bone resorption by osteoclasts and bone formation by osteoblasts is required for bone homeostasis. In a pathological context, enhanced osteoclastic bone resorption leads to a reduction of bone mass, as seen in osteoporosis, while defects in either osteoclast differentiation or function result in an abnormal accumulation of bone, as observed in osteopetrosis. Genetic studies of patients with osteopetrosis and naturally occurring or targeted gene knockout in rodent osteopetrotic models have unveiled important regulatory mechanisms of osteoclastic bone resorption. Disruption of genes such as c-src, ¿3 integrin subunit, cathepsin K, TCIRG1 (encoding a3 subunit of vacuolar H+-ATPase), CLCN7 (encoding clc-7 chloride channel) and OSTM1 (grey lethal) disrupts osteoclast function. Although mutations in TCIRG1 and CLCN7 are most abundant in human osteopetrosis patients, the involved genes have not been identified in about 25% of cases. Recently, PLEKHM1 was identified as a gene mutated in the osteopetrotic ia/ia (incisors absent) rat and a subset of patients with osteopetrosis. Osteoclasts from ia/ia rats are dysfunctional due to impaired ruffled border formation and secretion. However, the detailed mechanisms by which PLEKHM1 regulates osteoclast function remain unknown. We find that 1) expression of PLEKHM1 increases during osteoclast differentiation 2) knockdown of PLEKHM1 by shRNA inhibits cathepsin K secretion and osteoclast function, without a decrease in osteoclast formation and 3) plekhm1 binds to TRAF6 and rab7 in mature osteoclasts, two proteins critical for the cell's capacity to resorb bone. Thus, we hypothesize that plekhm1 regulates osteoclast function, at least in part, through interaction with TRAF6 and rab7. Since we have developed a retroviral system by which we can knockdown an endogenous gene and simultaneously rescue it with an RNAi-resistant mutant, we are in a position to explore the role of plekhm1 in osteoclast function by performing structure/function analysis. Therefore, our specific aims are to identify the domain or motif in plekhm1 essential for 1) plekhm11/TRAF6 interaction and osteoclast function and 2) rab7 binding and osteoclast function. The osteoclast, a cell found exclusively in bone, is responsible for dissolution of this important tissue. The aging American population continues to lose bone because of osteoclast activity, leading to major public health problems that currently cost in excess of $15 billion dollars annually. We have identified a novel protein in osteoclasts which contributes to their bone resorbing activity. We plan to determine how this new protein regulates the osteoclast, raising the possibility that we will be able to develop a new drug that prevents bone loss.

描述(申请人提供)：破骨细胞的骨吸收和成骨细胞的骨形成之间的紧密平衡是骨稳态所必需的。在病理学背景下，破骨细胞性骨吸收增强导致骨量减少，如骨质疏松，而破骨细胞分化或功能缺陷导致骨异常堆积，如骨质疏松。对骨质疏松患者和啮齿动物骨质疏松模型中自然发生或靶向基因敲除的遗传学研究揭示了破骨细胞性骨吸收的重要调节机制。C-src、整合素β3亚基、组织蛋白K、TCIRG1(编码空泡H+-ATPase的A3亚基)、CLCN7(编码ClC-7氯离子通道)和OSTM1(灰色致死)等基因的缺失破坏了破骨细胞的功能。尽管TCIRG1和CLCN7突变在人类骨化症患者中最为丰富，但约25%的病例尚未确定涉及的基因。最近，PLEKHM1基因被鉴定为一种突变基因，存在于骨质疏松性IA/IA(门牙缺失)大鼠和部分骨质疏松患者中。IA/IA大鼠的破骨细胞由于褶皱边缘的形成和分泌受损而功能失调。然而，PLEKHM1调节破骨细胞功能的详细机制仍不清楚。我们发现：1)在破骨细胞分化过程中，PLEKHM1的表达增加；2)shRNA下调PLEKHM1可抑制组织蛋白酶K的分泌和破骨细胞的功能，但不影响破骨细胞的形成；3)在成熟的破骨细胞中，PLEKHM1与TRAF6和Rab7结合，这两种蛋白质对细胞吸收骨的能力至关重要。因此，我们假设plekhm1至少部分地通过与TRAF6和Rab7的相互作用来调节破骨细胞的功能。由于我们已经开发了一种逆转录病毒系统，通过它我们可以敲除内源基因，同时用RNAi抗性突变体拯救它，因此我们能够通过结构/功能分析来探索plekhm1在破骨细胞功能中的作用。因此，我们的特定目标是确定plekhm1中对1)plekhm11/TRAF6相互作用和破骨细胞功能以及2)rab7结合和破骨细胞功能所必需的结构域或基序。破骨细胞是一种专门存在于骨骼中的细胞，负责分解这一重要组织。由于破骨细胞的活动，美国老龄化的人口继续失去骨骼，导致了重大的公共健康问题，目前每年的成本超过150亿美元。我们已经在破骨细胞中发现了一种新的蛋白质，它有助于其骨吸收活动。我们计划确定这种新的蛋白质是如何调节破骨细胞的，这增加了我们能够开发一种防止骨丢失的新药的可能性。