Yi X Y, Wayner E A, Kim Y, Fish A J
Department of Pediatrics, University of Minnesota, Minneapolis 55455, USA.
Cell Adhes Commun. 1998 Mar;5(3):237-48. doi: 10.3109/15419069809040294.
Entactin is an extracellular matrix glycoprotein which binds to laminin and is found in most renal basement membranes and in the glomerular mesangial matrix. In the present study, we have characterized specific integrin receptors on cultured human mesangial cells (CHMC) responsible for adhesion to native entactin. The integrin receptors alpha 2 beta 1, alpha 3 beta 1, alpha 5 beta 1, alpha v beta 3, alpha v beta 5, and alpha 6 complexed with either beta 1 or beta 4 could be immune precipitated from detergent extracts of metabolically labeled CHMC. Adhesion assays with inhibitory anti integrin monoclonal antibodies (mab) demonstrated that CHMC use both alpha v beta 3 and a beta 1-containing integrin to bind surfaces coated with native entactin. Optimal binding of CHMC to native entactin required the participation of cations. Using wild type and mutant recombinant entactin fragments, the binding site for the alpha v beta 3 receptor was localized to the RGD sequence on the rod or E domain of entactin. CHMC adhesion to mutant full length recombinant entactin ligands lacking the E domain RGD sequence confirmed the presence of ligand binding site(s) for beta 1 integrin receptor(s). Differences in CHMC binding characteristics to recombinant and full length entactin compared to native bovine basement membrane entactin were observed. This suggests that tertiary molecular structure may contribute to entactin ligand binding properties. Primary amino acid residue sequences and tertiary structure of entactin may play roles in forming functional cell attachment sites in native basement membrane entactin.
巢蛋白是一种细胞外基质糖蛋白，它与层粘连蛋白结合，存在于大多数肾基底膜和肾小球系膜基质中。在本研究中，我们已鉴定出培养的人系膜细胞（CHMC）上负责与天然巢蛋白黏附的特定整合素受体。整合素受体α2β1、α3β1、α5β1、αvβ3、αvβ5以及与β1或β4复合的α6可从经代谢标记的CHMC的去污剂提取物中免疫沉淀出来。用抑制性抗整合素单克隆抗体（mab）进行的黏附试验表明，CHMC利用αvβ3和含β1的整合素结合包被有天然巢蛋白的表面。CHMC与天然巢蛋白的最佳结合需要阳离子的参与。利用野生型和突变型重组巢蛋白片段，αvβ3受体的结合位点定位于巢蛋白杆状或E结构域上的RGD序列。CHMC对缺乏E结构域RGD序列的突变全长重组巢蛋白配体的黏附证实了β1整合素受体配体结合位点的存在。观察到与天然牛基底膜巢蛋白相比，CHMC对重组巢蛋白和全长巢蛋白的结合特性存在差异。这表明三级分子结构可能有助于巢蛋白的配体结合特性。巢蛋白的一级氨基酸残基序列和三级结构可能在天然基底膜巢蛋白中形成功能性细胞附着位点方面发挥作用。
