Plastin 3: Unravelling a novel pathomechanism for osteoporosis

骨质疏松症是老年人最常见的十种疾病之一，尤其影响绝经后的女性。骨质疏松症会导致骨折，尤其是在上肢、股骨颈和椎体。早期骨质疏松症合并骨折和成骨不全(OI)是由软骨形成和组装、骨生成或吸收相关的各种基因突变引起的单基因遗传性疾病，发病年龄和严重程度具有很高的变异性。在与Gerard Pals(阿姆斯特丹)和Carola Zillikens(鹿特丹)团队的富有成效的合作中，我们成功地在早期骨质疏松症合并骨折的男性和患有轻度骨质疏松症的女性中识别了X-连锁纤溶酶3基因(PLS3)的家族性致病变异。此外，在鹿特丹的一个大型队列中，发现了与PLS3基因SNP最显著的关联，这与女性的低骨密度和骨质疏松症有关。PLS3是一种钙离子依赖的F-肌动蛋白结合蛋白，在细胞迁移、内吞作用和所有依赖F-肌动蛋白的细胞过程中发挥重要作用。我们已经证明，在斑马鱼中，吗啡介导的PLS3基因敲除会导致头面部骨骼和肌肉结构的戏剧性畸形。通过联合注射PLS3RNA，这些患者被完全挽救(van Dijk等人。N Engl J Med 2013)。在另一个项目中，我们表明在女性中，PLS3的过度表达是脊髓性肌萎缩症(SMA)的保护性修饰物，SMA是一种常见的神经肌肉疾病(Oprea等人。科学，2008)。为了揭示保护机制，我们制造了一只条件性过表达PLS3的小鼠，我们证明了神经肌肉连接水平上的所有F-肌动蛋白绑定过程都得到了恢复(Ackermann等人Hum Mol Genet 2013)。该项目产生的初步数据显示，过度表达PLS3的小鼠具有更厚的皮质和骨小梁结构。PLS3的过度表达可以在大约5%的普通人群中发现，并且可能与预防骨质疏松或OI有关。在这个应用的背景下，我们的目标是通过使用条件基因敲除小鼠来解开PLS3缺失的致病机制。我们将产生无处不在或细胞类型缺乏PLS3的小鼠--特别是通过耗尽成骨细胞/骨细胞或骨骼肌中的PLS3。我们将进行一系列的体外和体外的形态学、组织学、细胞学、分子和生化实验，以确定PLS3缺失导致骨质疏松症的潜在发病机制。我们确定了10个新的与PLS3相互作用的伙伴，它们是解开分子机制的宝贵资源。了解骨质疏松症的分子发病机制将为未来的治疗开辟新的途径。

Osteoporosis represents one of the ten most frequently occurring diseases in elderly people, particularly affecting women after menopause. Osteoporosis causes fractures especially in the upper limbs, femoral neck and vertebral bodies. Early osteoporosis with fractures and osteogenesis imperfecta (OI) are monogenic inherited diseases with a high variability in age of onset and severity, caused by mutations in various genes involved in cartilage formation and assembly, bone generation or resorption. In a productive collaborative work with the groups of Gerard Pals (Amsterdam) and Carola Zillikens (Rotterdam), we succeeded in identifying familial pathogenic variants in the X-linked Plastin 3 gene (PLS3) in men with early osteoporosis with fractures and women with mild osteoporosis. Furthermore, in a large Rotterdam cohort the most significant association with a SNP in PLS3 was identified, which correlates with low bone mineral density and osteoporosis in women. PLS3 is a Ca2+-dependent F-actin bundling protein with an essential role in cell migration, endocytosis und all F-actin-dependent cellular processes. We have shown that morpholino-mediated pls3 knockdown in zebrafish causes dramatic deformities in craniofacial skeletal and muscle structures. These were fully rescued by co-injection of PLS3 RNA (van Dijk et al. N Engl J Med 2013). In another project we showed that in women overexpression of PLS3 is a protective modifier for spinal muscular atrophy (SMA), a frequently occurring neuromuscular disease (Oprea et al. Science 2008). To unravel the protective mechanism, we generated a conditional PLS3 overexpressing mouse and we showed that all F-actin bundling processes at the neuromuscular junction level are restored (Ackermann et al Hum Mol Genet 2013). Preliminary data generated in this project showed that mice overexpressing PLS3 have thicker cortical and trabecular bone structures. PLS3 overexpression can be found in about 5% of the general population and might correlate with protection against osteoporosis or OI. In the context of this application, we aim at unravelling the pathomechanism underlying PLS3 depletion by using conditional knock-out mice. We will generate mice lacking Pls3 either ubiquitously or cell-type -specifically by depleting Pls3 in osteoblasts/osteocytes or in skeletal muscle. A full battery of morphological, histological, cellular, molecular and biochemical in vitro and ex vivo experiments will be carried out to determine the underlying pathomechanism of osteoporosis caused by loss of PLS3. We identified 10 novel PLS3-interacting partners, which represent valuable resources to unravel the molecular mechanism. Understanding the molecular pathomechanism of osteoporosis will open new avenues for future therapies.