A critical role for PLD1 in osteoclast fusion

摘要 细胞-细胞融合是一种对几个发育和生理过程至关重要的细胞事件 包括骨骼动态平衡。骨平衡是由成骨细胞和骨吸收维持的 破骨细胞。破骨细胞的融合对破骨细胞的激活和功能至关重要。高架 破骨细胞生成和骨吸收导致骨质疏松症，这是一个严重的健康问题，其特征是 衰老和病理条件下的骨重建失衡。骨的抗吸收治疗现状 针对早期破骨细胞分化和/或活性的丢失，一直受到低应答率或 副作用。最近的研究表明，靶向分子控制破骨细胞融合可能是一种更好的抗 骨丢失的吸收治疗策略。然而，人们对其分子机制知之甚少。 潜在的破骨细胞融合。我们最近发现磷脂酶D_1(PLD_1)，通过产生 信号转导的磷脂磷脂酸(PA)在破骨细胞融合中起着关键作用。我们的体外实验 显示PLD1活性在破骨细胞形成过程中被激活。PLD1的抑制或基因缺失抑制了 单核破骨细胞与多核破骨细胞的融合。PLD1通过促进破骨细胞融合 调节融合突触突起膜结构的形成。通过两个PA- 结合蛋白质筛选和RNA-Seq，我们已经确定了PLD1调节的一个潜在机制 破骨细胞融合。最后，编码PLD1的Pld1基因的全局缺失可以保护小鼠免受骨质丢失的影响。 基于这些数据，我们假设PLD1产生的PA在破骨细胞融合中起关键作用 调节突触融合突触的形成。我们将使用以下内容来验证我们的假设 明确的目标。在目标1中，我们将证明PLD1生成的PA的时空产生是 对破骨细胞融合至关重要。在目标2中，我们将阐明PLD1协调肌动蛋白的机制 破骨细胞融合过程中的细胞骨架重组和膜重塑。在目标3中，我们将定义 利用基因敲除技术研究破骨细胞PLD1在生理和病理条件下骨稳态中的作用 老鼠模型。这个项目的目标是阐明pld1在破骨细胞融合和骨中的作用。 动态平衡，并确定新的和更有效的治疗骨质疏松症和其他骨骼疾病的靶点。

Abstract Cell-cell fusion is a cellular event that is critical to several developmental and physiological processes including bone homeostasis. Bone homeostasis is maintained by bone-forming osteoblasts and bone-resorbing osteoclasts. The fusion of osteoclasts is critical for osteoclast activation and function. Elevated osteoclastogenesis and bone resorption leads to osteoporosis that is a severe health problem characterized by imbalanced bone remodeling in aging and pathological conditions. Current anti-resorptive treatments for bone loss, which target early osteoclast differentiation and/or viability, have been limited by low response rates or side effects. Recent studies suggest that targeting molecules controling osteoclast fusion may be a better anti- resorptive therapeutic strategy for bone loss. However, very little is known about the molecular mechanisms underlying osteoclast fusion. We recently found that phospholipase D1 (PLD1), through generating the signaling phospholipid phosphatidic acid (PA), plays a critical role in osteoclast fusion. Our In vitro experiments show that PLD1 activity is activated during osteoclastogenesis. Inhibition or genetic deletion of PLD1 inhibit the fusion of mononucleated osteoclasts to multinucleated osteoclasts. PLD1 promotes osteoclast fusion through regulating the formation of the protrusive membrane structure at fusogenic synapses. Through both a PA- binding protein screen and RNA-Seq, we have identified a potential mechanism through which PLD1 regulates osteoclast fusion. Finally, global deletion of Pld1, the gene encoding PLD1, protects mice from bone loss. Based on these data, we hypothesize PLD1-generated PA plays a critical role in osteoclast fusion through regulating the formation of the protrusive fusogenic synapse. We will test our hypothesis with the following specific aims. In Aim 1, we will demonstrate that the spatiotemporal production of PLD1-genereated PA is critical for osteoclast fusion. In Aim 2, we will elucidate the mechanisms through which PLD1 coordinates actin cytoskeletal reorganization and membrane remodeling during osteoclast fusion. In Aim 3, we will define the role of osteoclast PLD1 in bone homeostasis in both physiological and pathological conditions using knockout mouse models. The goals of this project are to elucidate the role of PLD1 in osteoclast fusion and bone homeostasis and identify new and more effective therapeutic targets for osteoporosis and other bone diseases.