Laboratory of Microbiology and Immunology, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka City, Shizuoka 422-8526, Japan.
Laboratory of Plant Molecular Improvement, Graduate Division of Nutritional and Environmental Sciences, University of Shizuoka, Shizuoka City, Shizuoka 422-8526, Japan.
Sci Rep. 2017 Apr 3;7:45843. doi: 10.1038/srep45843.
Shiga toxin is a major virulence factor of food-poisoning caused by Escherichia coli such as O157:H7. Secretory immunoglobulin (Ig) A (SIgA) is supposed to prevent infection of the mucosal surface and is a candidate agent for oral immunotherapy. We previously established a recombinant monoclonal antibody (mAb) consisting of variable regions from a mouse IgG mAb specific for the binding subunit of Shiga toxin 1 (Stx1) and the Fc region of mouse IgA. Here we produced a secretory form of the recombinant IgA (S-hyIgA) with transgenic Arabidopsis thaliana plant. All the S-hyIgA cDNAs (heavy, light, J chain and secretory component) were expressed under the control of a bidirectional promoter of a chlorophyll a/b-binding protein of A. thaliana without using a viral promoter. The plant-based S-hyIgA exhibited antigen binding, and was modified with plant-specific N-linked sugar chains. The Ig heavy chain and secretory components were observed in an intracellular protein body-like structure of the transgenic leaves on immuno-electron microscopy. An extract of the transgenic leaves neutralized the cytotoxicity of Stx1 toward butyrate-treated Caco-2 cells, a human colon carcinoma cell line. These results will contribute to the development of edible therapeutic antibodies such as those for the treatment of mucosal infection.
志贺毒素是由大肠杆菌（如 O157：H7）引起的食物中毒的主要毒力因子。分泌型免疫球蛋白（Ig）A（SIgA）被认为可以预防黏膜表面感染，是口服免疫治疗的候选药物。我们之前建立了一种由针对志贺毒素 1（Stx1）结合亚基的小鼠 IgG mAb 的可变区和小鼠 IgA 的 Fc 区组成的重组单克隆抗体（mAb）。在这里，我们使用转基因拟南芥植物生产了重组 IgA（S-hyIgA）的分泌形式。所有 S-hyIgA cDNA（重链、轻链、J 链和分泌成分）均在拟南芥叶绿素 a/b 结合蛋白的双向启动子控制下表达，而不使用病毒启动子。基于植物的 S-hyIgA 表现出抗原结合，并经过植物特异性 N-连接糖链修饰。在免疫电子显微镜下，可在转基因叶片的细胞内蛋白体样结构中观察到 Ig 重链和分泌成分。转基因叶片提取物中和了 Stx1 对丁酸盐处理的人结肠癌细胞系 Caco-2 细胞的细胞毒性。这些结果将有助于开发可食用的治疗性抗体，例如用于治疗黏膜感染的抗体。
