Mechanisms of Blister Formation by Staphylococcal Toxins

剥脱毒素A(ETA)由金黄色葡萄球菌产生，可引起葡萄球菌性烫伤皮肤综合征(SSSS)及其更局限性的大疱性脓疱病。ETA的晶体结构表明，它是一种丝氨酸蛋白酶，当ETA与特定受体结合时，该酶具有不活跃的催化部位而被激活。在叶状天疱疮中，导致DSG 1功能障碍的自身抗体会导致水泡，与ETA在小鼠和人的浅层表皮中引起的水泡相同。因此，我们假设DSG1特异性地结合并激活ETA，ETA进而切割结合的DSG1，导致水泡的形成。我们认为，另一种引起大疱性脓疱病和SSSS的葡萄球菌毒素--脱落毒素B(ETB)也被DSG-1激活和裂解。最后，我们推测ETA与DSG-1结合和/或ETA裂解DSG-1可能引起对DSG-1的自身免疫反应，从而提示了PF患者产生自身抗体的机制。我们已经证明了ETA切割DSG1。特定的目标1将通过确定切割是否依赖于DSG1的构象来表征这种切割，并通过定义切割位置和切割所需的DSG1结构域来表征这种切割。目标2将表征ETA与DSG 1的结合，并定义每个必需的结构域。目的利用DSG3基因敲除小鼠和白藜芦醇-DSG3转基因小鼠，确定DSG3的代偿能否弥补ETA诱导的DSG1功能丧失，从而解释水泡定位的部位。目的4包含旨在确定ETA切割DSG 1的动力学的研究。目标5将扩大以前目标的成果，以纳入ETB的行动机制。最终目标将确定大疱性脓疱病和SSSS患者是否产生针对DSG 1的抗体反应，叶型天疱疮患者是否对ETA和ETB有增强的免疫反应，以及注射ETA的小鼠是否对DSG 1产生免疫反应。这些研究将提供关于一种非常常见的疾病-大疱性脓疱病的分子病理生理学的见解，并首次确定一种组织特异性自身免疫性疾病天疱疮的潜在触发或加重因素。

Exfoliative toxin A (ETA), produced by Staphylococcus aureus, causes staphylococcal scalded skin syndrome (SSSS) and its more localized form, bullous impetigo. The crystal structure of ETA suggests that it is a serine protease with an inactive catalytic site which becomes activated when ETA binds a specific receptor. In pemphigus foliaceus autoantibodies that cause dysfunction of Dsg 1 cause blisters identical to those caused by ETA in the superficial epidermis of mouse and man. Therefore, we hypothesize that Dsg 1 specifically binds and activates ETA, which in turn cleaves the bound Dsg 1, resulting in blister formation. We propose that another staphylococcal toxin, exfoliative toxin B (ETB), that also causes bullous impetigo and SSSS, is also activated by, and cleaves, Dsg 1. Finally, we hypothesize that binding of ETA to Dsg 1 and/or cleavage of Dsg 1 by ETA might elicit an autoimmune response against Dsg 1, thus suggesting a mechanism for autoantibody production in PF patients. We have shown that ETA cleaves Dsg 1. Specific aim 1 will characterize this cleavage by determining if cleavage is dependent on Dsg 1 conformation, and by defining the site of cleavage and the domains of Dsg 1 needed for cleavage. Aim 2 will characterize binding of ETA to Dsg 1, and define the domains of each necessary. Aim 3 will determine, using Dsg 3 knockout and involucrin-Dsg 3 transgenic mice, if compensation by Dsg 3 can compensate for ETA-induced loss of function of Dsg 1, thereby explaining the sites of blister localization. Aim 4 contains studies designed to define the kinetics of Dsg 1 cleavage by ETA. Aim 5 will extend the results of the previous aims to include the mechanisms of action of ETB. The final aim will determine if patients with bullous impetigo and SSSS develop an antibody response against Dsg 1, if patients with pemphigus foliaceus have an enhanced immune response against ETA and ETB, and if mice injected with ETA develop an immune response against Dsg 1. These studies will provide insight regarding the molecular pathophysiology of a very common disease, bullous impetigo, and, for the first time, identify a potential trigger or exacerbating factor in a tissue-specific autoimmune disease, pemphigus.