PDF(Pubmed)
Abstract：
Mutations in PLEKHM1 cause osteopetrosis in humans and rats. The germline and osteoclast conditional deletions of Plekhm1 gene in mice lead to defective osteoclast bone resorption and increased trabecular bone mass without overt abnormalities in other organs. As an adaptor protein, pleckstrin homology and RUN domain containing M1 (PLEKHM1) interacts with the key lysosome regulator small GTPase RAB7 via its C-terminal RUBICON homologous (RH) domain. In this study, we have conducted a structural-functional study of the PLEKHM1 RH domain and RAB7 interaction in osteoclasts in vitro. The single mutations of the key residues in the Plekhm1 RH predicted from the crystal structure of the RUBICON RH domain and RAB7 interface failed to disrupt the Plekhm1-Rab7 binding, lysosome trafficking, and bone resorption. The compound alanine mutations at Y949-R954 and L1011-I1018 regions decreased Plekhm1 protein stability and Rab7-binding, respectively, thereby attenuated lysosome trafficking and bone resorption in osteoclasts. In contrast, the compound alanine mutations at R1060-Q1068 region were dispensable for Rab7-binding and Plekhm1 function in osteoclasts. These results indicate that the regions spanning Y949-R954 and L1011-I1018 of Plekhm1 RH domain are functionally important for Plekhm1 in osteoclasts and offer the therapeutic targets for blocking bone resorption in treatment of osteoporosis and other metabolic bone diseases.
摘要：
PLEKHM1中的突变在人和大鼠中引起骨硬化。小鼠Plekhm1基因的种系和破骨细胞条件缺失导致破骨细胞骨吸收缺陷和小梁骨量增加，而其他器官没有明显异常。作为一种衔接蛋白，pleckstrin同源性和含有M1的RUN域（PLEKHM1）通过其C端RUBICON同源（RH）域与关键溶酶体调节因子小GTP酶RAB7相互作用。在这项研究中,我们对体外破骨细胞中PLEKHM1RH结构域和RAB7相互作用进行了结构-功能研究.从RUBICONRH结构域和RAB7界面的晶体结构预测的Plekhm1RH中关键残基的单突变未能破坏Plekhm1-Rab7结合，溶酶体贩运,和骨吸收。在Y949-R954和L1011-I1018区域的复合丙氨酸突变降低了Plekhm1蛋白的稳定性和Rab7结合，分别,从而减弱溶酶体运输和破骨细胞中的骨吸收。相比之下,R1060-Q1068区域的复合丙氨酸突变对于破骨细胞中的Rab7结合和Plekhm1功能是不必要的。这些结果表明，跨Plekhm1RH结构域的Y949-R954和L1011-I1018的区域对于破骨细胞中的Plekhm1在功能上是重要的，并且为在骨质疏松症和其他代谢性骨疾病的治疗中阻断骨吸收提供了治疗靶标。
