Department of Orthopaedic Surgery, University of California-Los Angeles, 615 Charles E. Young Drive South, BSRB 512, Los Angeles, CA 90095, United States.
Department of Orthopaedic Surgery, University of California-Los Angeles, 615 Charles E. Young Drive South, BSRB 512, Los Angeles, CA 90095, United States; Laboratory of Bioengineering and Tissue Regeneration-LABRET, Department of Cell Biology, Genetics and Physiology, University of Málaga, IBIMA, Málaga 29071, Spain.
EBioMedicine. 2020 Dec;62:103075. doi: 10.1016/j.ebiom.2020.103075. Epub 2020 Nov 23.
Beyond its structural role in the skeleton, the extracellular matrix (ECM), particularly basement membrane proteins, facilitates communication with intracellular signaling pathways and cell to cell interactions to control differentiation, proliferation, migration and survival. Alterations in extracellular proteins cause a number of skeletal disorders, yet the consequences of an abnormal ECM on cellular communication remains less well understood METHODS: Clinical and radiographic examinations defined the phenotype in this unappreciated bent bone skeletal disorder. Exome analysis identified the genetic alteration, confirmed by Sanger sequencing. Quantitative PCR, western blot analyses, immunohistochemistry, luciferase assay for WNT signaling were employed to determine RNA, proteins levels and localization, and dissect out the underlying cell signaling abnormalities. Migration and wound healing assays examined cell migration properties.
This bent bone dysplasia resulted from biallelic mutations in LAMA5, the gene encoding the alpha-5 laminin basement membrane protein. This finding uncovered a mechanism of disease driven by ECM-cell interactions between alpha-5-containing laminins, and integrin-mediated focal adhesion signaling, particularly in cartilage. Loss of LAMA5 altered β1 integrin signaling through the non-canonical kinase PYK2 and the skeletal enriched SRC kinase, FYN. Loss of LAMA5 negatively impacted the actin cytoskeleton, vinculin localization, and WNT signaling.
This newly described mechanism revealed a LAMA5-β1 Integrin-PYK2-FYN focal adhesion complex that regulates skeletogenesis, impacted WNT signaling and, when dysregulated, produced a distinct skeletal disorder.
Supported by NIH awards R01 AR066124, R01 DE019567, R01 HD070394, and U54HG006493, and Czech Republic grants INTER-ACTION LTAUSA19030, V18-08-00567 and GA19-20123S.
细胞外基质（ECM）除了在骨骼中具有结构作用外，还特别有助于与细胞内信号通路和细胞间相互作用进行交流，从而控制分化、增殖、迁移和存活。细胞外蛋白的改变会导致许多骨骼疾病，但 ECM 对细胞通讯的影响仍然知之甚少。
临床和影像学检查定义了这种未被认识的弯曲骨骨骼疾病的表型。外显子组分析确定了遗传改变，并通过 Sanger 测序进行了确认。定量 PCR、western blot 分析、免疫组织化学、WNT 信号的荧光素酶测定用于确定 RNA 和蛋白质水平和定位，并剖析潜在的细胞信号异常。迁移和伤口愈合试验检查了细胞迁移特性。
这种弯曲骨发育不良是由编码α-5 层粘连蛋白基底膜蛋白的 LAMA5 基因的双等位基因突变引起的。这一发现揭示了一种由 ECM-细胞相互作用驱动的疾病机制，特别是在软骨中，α-5 层粘连蛋白与整合素介导的焦点粘附信号之间存在相互作用。LAMA5 的缺失通过非典型激酶 PYK2 和骨骼丰富的 SRC 激酶 FYN 改变了β1 整合素信号。LAMA5 的缺失对肌动蛋白细胞骨架、粘着斑蛋白 vinculin 的定位和 WNT 信号产生负面影响。
新描述的机制揭示了 LAMA5-β1 整合素-PYK2-FYN 焦点粘附复合物，该复合物调节骨骼发生，影响 WNT 信号，当失调时会产生独特的骨骼疾病。
由 NIH 授予的 R01 AR066124、R01 DE019567、R01 HD070394 和 U54HG006493，以及捷克共和国授予的 INTER-ACTION LTAUSA19030、V18-08-00567 和 GA19-20123S 资助。
Zotero 插件
