Department of Microbiology, School of Molecular Biosciences, Washington State University, Pullman, Washington, USA.
J Bacteriol. 2010 Jan;192(1):68-76. doi: 10.1128/JB.00969-09.
Campylobacter jejuni is one of the most frequent bacterial causes of food-borne gastrointestinal disease in developed countries. Previous work indicates that the binding of C. jejuni to human intestinal cells is crucial for host colonization and disease. Fibronectin (Fn), a major constituent of the extracellular matrix, is a approximately 250-kDa glycoprotein present at regions of cell-to-cell contact in the intestinal epithelium. Fn is composed of three types of repeating units: type I (approximately 45 amino acids), type II (approximately 60 amino acids), and type III (approximately 90 amino acids). The deduced amino acid sequence of C. jejuni flpA (Cj1279c) contains at least three Fn type III domains. Based on the presence of the Fn type III domains, we hypothesized that FlpA contributes to the binding of C. jejuni to human INT 407 epithelial cells and Fn. We assessed the contribution of FlpA in C. jejuni binding to host cells by in vitro adherence assays with a C. jejuni wild-type strain and a C. jejuni flpA mutant and binding of purified FlpA protein to Fn by enzyme-linked immunosorbent assay (ELISA). Adherence assays revealed the binding of the C. jejuni flpA mutant to INT 407 epithelial cells was significantly reduced compared with that for a wild-type strain. In addition, rabbit polyclonal serum generated against FlpA blocked C. jejuni adherence to INT 407 cells in a concentration-dependent manner. Binding of FlpA to Fn was found to be dose dependent and saturable by ELISA, demonstrating the specificity of the interaction. Based on these data, we conclude that FlpA mediates C. jejuni attachment to host epithelial cells via Fn binding.
空肠弯曲菌是发达国家中最常见的食源性胃肠道疾病细菌病原体之一。先前的工作表明，空肠弯曲菌与人肠道细胞的结合对于宿主定植和疾病的发生至关重要。纤连蛋白（Fn）是细胞外基质的主要成分之一，是一种约 250kDa 的糖蛋白，存在于肠道上皮细胞的细胞间接触区域。Fn 由三种重复单元组成：I 型（约 45 个氨基酸）、II 型（约 60 个氨基酸）和 III 型（约 90 个氨基酸）。空肠弯曲菌 flpA（Cj1279c）的推导氨基酸序列至少包含三个 Fn III 结构域。基于 Fn III 结构域的存在，我们假设 FlpA 有助于空肠弯曲菌与人 INT 407 上皮细胞和 Fn 的结合。我们通过体外粘附试验评估了 FlpA 在空肠弯曲菌与宿主细胞结合中的作用，该试验使用空肠弯曲菌野生型菌株和 flpA 突变体以及纯化的 FlpA 蛋白与 Fn 的酶联免疫吸附试验（ELISA）。粘附试验显示，与野生型菌株相比，flpA 突变体与 INT 407 上皮细胞的结合显著减少。此外，针对 FlpA 产生的兔多克隆血清以浓度依赖的方式阻断了空肠弯曲菌对 INT 407 细胞的粘附。通过 ELISA 发现 FlpA 与 Fn 的结合呈剂量依赖性和饱和性，表明相互作用具有特异性。基于这些数据，我们得出结论，FlpA 通过与 Fn 结合介导空肠弯曲菌附着到宿主上皮细胞。
