Department of Biomedical Engineering, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio 44195, USA.
J Biol Chem. 2011 May 13;286(19):17156-67. doi: 10.1074/jbc.M111.231571. Epub 2011 Mar 14.
Autosomal recessive and autosomal dominant forms of Weill-Marchesani syndrome, an inherited connective tissue disorder, are caused by mutations in ADAMTS10 (encoding a secreted metalloprotease) and FBN1 (encoding fibrillin-1, which forms tissue microfibrils), respectively, yet they are clinically indistinguishable. This genetic connection prompted investigation of a potential functional relationship between ADAMTS10 and fibrillin-1. Specifically, fibrillin-1 was investigated as a potential ADAMTS10 binding partner and substrate, and the role of ADAMTS10 in influencing microfibril biogenesis was addressed. Using ligand affinity blotting and surface plasmon resonance, recombinant ADAMTS10 was found to bind to fibrillin-1 with a high degree of specificity and with high affinity. Two sites of ADAMTS10 binding to fibrillin-1 were identified, one toward the N terminus and another in the C-terminal half of fibrillin-1. Confocal microscopy and immunoelectron microscopy localized ADAMTS10 to fibrillin-1-containing microfibrils in human tissues. Furin-activated ADAMTS10 could cleave fibrillin-1, but innate resistance of ADAMTS10 zymogen to propeptide excision by furin was observed, suggesting that, unless activated, ADAMTS10 is an inefficient fibrillinase. To investigate the role of ADAMTS10 in microfibril biogenesis, fetal bovine nuchal ligament cells were cultured in the presence or absence of ADAMTS10. Exogenously added ADAMTS10 led to accelerated fibrillin-1 microfibril biogenesis. Conversely, fibroblasts obtained from a Weill-Marchesani syndrome patient with ADAMTS10 mutations deposited fibrillin-1 microfibrils sparsely compared with unaffected control cells. Taken together, these findings suggest that ADAMTS10 participates in microfibril biogenesis rather than in fibrillin-1 turnover.
常染色体隐性和常染色体显性遗传的 Marchesani 综合征（一种遗传性结缔组织疾病）分别由 ADAMTS10（编码一种分泌型金属蛋白酶）和 FBN1（编码纤维连接蛋白-1，后者形成组织微纤维）的突变引起，但它们在临床上无法区分。这种遗传联系促使人们研究 ADAMTS10 和纤维连接蛋白-1 之间潜在的功能关系。具体来说，纤维连接蛋白-1 被研究为 ADAMTS10 的潜在结合伴侣和底物，以及 ADAMTS10 影响微纤维发生的作用。通过配体亲和印迹和表面等离子体共振，发现重组 ADAMTS10 与纤维连接蛋白-1 具有高度特异性和高亲和力结合。鉴定了 ADAMTS10 与纤维连接蛋白-1 结合的两个位点，一个在纤维连接蛋白-1 的 N 端，另一个在纤维连接蛋白-1 的 C 端。共聚焦显微镜和免疫电子显微镜将 ADAMTS10 定位在人类组织中含有纤维连接蛋白-1 的微纤维上。激活的 Furin 可切割纤维连接蛋白-1，但 ADAMTS10 酶原对 Furin 前肽切割的固有抗性被观察到，这表明，除非被激活，否则 ADAMTS10 不是一种有效的纤维连接蛋白酶。为了研究 ADAMTS10 在微纤维发生中的作用，在存在或不存在 ADAMTS10 的情况下培养胎牛颈韧带细胞。外源性添加 ADAMTS10 导致纤维连接蛋白-1 微纤维发生加速。相反，与未受影响的对照细胞相比，从患有 ADAMTS10 突变的 Marchesani 综合征患者获得的成纤维细胞稀疏地沉积纤维连接蛋白-1 微纤维。总之，这些发现表明 ADAMTS10 参与微纤维发生，而不是纤维连接蛋白-1 周转。
