Identification of a domain within the multifunctional Vibrio cholerae RTX toxin that covalently cross-links actin
Identification of a domain within the multifunctional Vibrio cholerae RTX toxin that covalently cross-links actin
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DOI：
10.1073/pnas.0401104101
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发表时间：
2004-06-29
影响因子：
11.1
通讯作者：
Satchell, KJF
中科院分区：
文献类型：
--
作者：
Sheahan, KL;Cordero, CL;Satchell, KJF
革兰氏阴性病原体霍乱弧菌通过输出肠毒素引起腹泻病。霍乱弧菌RTX毒素先前通过其使人喉上皮（HEp-2）细胞变圆的能力被鉴定和表征。进一步的研究确定，细胞变圆是由肌动蛋白应力纤维解聚引起的，通过共价肌动蛋白交联的独特机制。在这项研究中，我们确定了一个域内的全长RTX毒素，能够介导的交联反应时，瞬时表达在真核细胞内。肌动蛋白交联结构域（ACD）的结构/功能分析表明，一个412-aa，或47.8-kDa的区域是必不可少的交联活性。当该结构域从全长毒素基因中删除时，肌动蛋白交联而不是细胞变圆被消除，表明该毒素具有多种可分离的活性。ACD与来自霍乱弧菌的假设蛋白VC 1416共享59%的氨基酸同一性，并且VC 1416的C-末端结构域的瞬时表达也导致真核细胞中的肌动蛋白交联。与Rhs样元件连接的第二个ACD的存在表明，霍乱弧菌通过水平基因转移获得了该结构域，并且ACD通过霍乱弧菌进化过程中的基因复制插入RTX毒素中。
The Gram-negative pathogen Vibrio cholerae causes diarrheal disease through the export of enterotoxins. The V. cholerae RTX toxin was previously identified and characterized by its ability to round human laryngeal epithelial (HEp-2) cells. Further investigation determined that cell rounding is caused by the depolymerization of actin stress fibers, through the unique mechanism of covalent actin cross-linking. In this study, we identify a domain within the full-length RTX toxin that is capable of mediating the cross-linking reaction when transiently expressed within eukaryotic cells. A structure/function analysis of the actin cross-linking domain (ACD) reveals that a 412-aa, or a 47.8-kDa, region is essential for cross-linking activity. When this domain is deleted from the full-length toxin gene, actin cross-linking, but not cell rounding, is eliminated, indicating that this toxin carries multiple dissociable activities. The ACD shares 59% amino acid identity with a hypothetical protein from V. cholerae, VC1416, and transient expression of the C-terminal domain of VC1416 also results in actin cross-linking in eukaryotic cells. The presence of this second ACD linked to an Rhs-like element suggests that V. cholerae acquired the domain by horizontal gene transfer and the ACD was inserted into the RTX toxin by gene duplication through the evolution of V. cholerae.