The Role of PLEKHM1 in Osteoclast Function

PLEKHM1 在破骨细胞功能中的作用

• 批准号: 7360975
• 负责人: HAIBO ZHAO
• 金额: $ 7.6万
• 依托单位: WASHINGTON UNIVERSITY
• 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 2011-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7360975
• 关键词: Aging; Albers-Schonberg disease; American; Binding; Bone Resorption; Cells; Chloride Channels; Defect; Disruption; 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; TNF receptor-associated factor 6; TRAF6 gene; Tissues; bone; bone loss; 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（由申请人提供）：骨稳态需要破骨细胞骨吸收和成骨细胞骨形成之间的紧密平衡。在病理背景下，破骨细胞骨吸收增强导致骨量减少，如骨质疏松症，而破骨细胞分化或功能缺陷导致骨异常积聚，如骨质疏松症。对骨质疏松患者的遗传学研究以及在啮齿动物骨质疏松模型中自然发生或靶向基因敲除，揭示了破骨细胞骨吸收的重要调控机制。c-src、¿3整合素亚基、组织蛋白酶K、TCIRG1（编码液泡H+- atp酶的a3亚基）、CLCN7（编码clc-7氯离子通道）和OSTM1（灰色致死）等基因的破坏会破坏破骨细胞的功能。尽管TCIRG1和CLCN7突变在人类骨质疏松症患者中最为丰富，但在约25%的病例中尚未发现相关基因。最近，PLEKHM1被鉴定为在骨质疏松大鼠（缺门牙）和骨质疏松患者亚群中发生突变的基因。ia/ia大鼠破骨细胞功能失调，原因是褶边形成和分泌受损。然而，PLEKHM1调控破骨细胞功能的详细机制尚不清楚。我们发现1)在破骨细胞分化过程中PLEKHM1的表达增加2)shRNA敲低PLEKHM1抑制cathepsin K分泌和破骨细胞功能，但不减少破骨细胞的形成3)PLEKHM1在成熟的破骨细胞中与TRAF6和rab7结合，这两种蛋白对细胞吸收骨的能力至关重要。因此，我们假设plekhm1至少部分通过与TRAF6和rab7的相互作用调节破骨细胞功能。由于我们已经开发了一种逆转录病毒系统，通过该系统我们可以敲除内源性基因，同时用rnai抗性突变体拯救它，我们可以通过进行结构/功能分析来探索plekhm1在破骨细胞功能中的作用。因此，我们的具体目标是确定plekhm1中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，一种只存在于骨中的细胞，负责分解这一重要组织。由于破骨细胞的活动，老龄化的美国人口继续失去骨质，导致重大的公共卫生问题，目前每年花费超过150亿美元。我们在破骨细胞中发现了一种有助于骨吸收活性的新蛋白。我们计划确定这种新蛋白质如何调节破骨细胞，从而提高我们开发一种防止骨质流失的新药的可能性。