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的骨吸收和成骨细胞的骨形成之间的紧密平衡。在病理背景下，破骨细胞的骨吸收增强导致骨量减少，如骨质疏松症中所见，而缺陷。破骨细胞分化或功能导致骨异常积累，正如在骨石症患者的遗传学研究和啮齿类动物中自然发生或靶向基因敲除中观察到的那样。骨石模型揭示了破骨细胞骨吸收的重要调节机制，例如 c-src、¿ 3 整合素亚基、组织蛋白酶 K、TCIRG1（编码空泡 H+-ATP 酶的 a3 亚基）、CLCN7（编码 clc-7 氯通道）和 OSTM1（灰色致死）会破坏破骨细胞功能，尽管 TCIRG1 和 CLCN7 的突变在人类骨硬化症中最为丰富。最近，大约25%的病例中所涉及的基因尚未被识别。 PLEKHM1 被鉴定为石骨症 ia/ia（门齿缺失）大鼠和一部分石骨症患者的基因，由于皱边形成和分泌受损，破骨细胞功能障碍。 PLEKHM1 调节破骨细胞功能仍然未知，1) PLEKHM1 的表达在破骨细胞分化过程中增加；2) PLEKHM1 的敲低。 shRNA 抑制组织蛋白酶 K 分泌和破骨细胞功能，但不减少破骨细胞形成，并且 3) plekhm1 与成熟破骨细胞中的 TRAF6 和 rab7 结合，这两种蛋白质对于细胞吸收骨的能力至关重要。因此，我们努力证明 plekhm1 调节破骨细胞功能。至少部分是通过与 TRAF6 和 rab7 的相互作用，因为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低内源基因并同时用 RNAi 抗性突变体来拯救它，我们能够通过进行结构/功能分析来探索 plekhm1 在破骨细胞功能中的作用，因此，我们的具体目标是识别 plekhm1 中对于 1) plekhm11 至关重要的结构域或基序。 /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 破骨细胞是一种专门存在于骨骼中的细胞，负责溶解这一重要组织。由于破骨细胞活性而导致骨质流失，导致目前每年造成的损失超过 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蛋白质如何调节。破骨细胞，提高了我们开发一种防止骨质流失的新药的可能性。