Mechanisms of Blister Formation by Staphylococcal Toxins

由金黄色葡萄球菌产生的剥脱毒素A （ETA）可引起葡萄球菌性烫伤皮肤综合征（SSSS）及其更局部的形式，大疱性脓疱病。ETA的晶体结构表明它是一种丝氨酸蛋白酶，具有非活性的催化位点，当ETA与特定受体结合时，催化位点被激活。在叶状天疱疮中，引起dsg1功能障碍的自身抗体在小鼠和人的浅表皮中引起与ETA相同的水泡。因此，我们假设Dsg 1特异性地结合并激活ETA， ETA反过来切割结合的Dsg 1，导致水疱形成。我们提出另一种葡萄球菌毒素，剥脱毒素B (ETB)，也引起大疱性脓疱病和SSSS，也被dsg1激活并裂解。最后，我们假设ETA与dsg1的结合和/或ETA对dsg1的切割可能引发针对dsg1的自身免疫反应，从而提示了PF患者自身抗体产生的机制。我们已经证明了ETA切割dsg1。具体目标1将通过确定切割是否依赖于dsg1构象，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将描述ETA与Dsg 1的结合，并定义每个必要的域。目的3将通过敲除dsg3和天花素- dsg3转基因小鼠，确定dsg3的补偿是否可以补偿eta诱导的dsg1功能丧失，从而解释水疱定位的位置。目的4包含旨在定义ETA裂解dsg1动力学的研究。目标5将扩展先前目标的结果，以包括ETB的行动机制。最终目的是确定大疱性脓疱疮和SSSS患者是否产生针对dsg1的抗体反应，叶状天疱疮患者是否对ETA和ETB有增强的免疫反应，以及注射ETA的小鼠是否产生针对dsg1的免疫反应。这些研究将为一种非常常见的疾病大疱疮的分子病理生理学提供见解，并首次确定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.