Bordetella cyclase toxin: effects on leukocytes and respiratory epithelial cells

描述(由申请人提供)：腺苷环化酶(AC)毒素是一种新的毒力因子，可用于波氏杆菌属的几个物种，包括百日咳杆菌(人类百日咳、百日咳)、副百日咳杆菌(动物和人类的不同菌株)和支气管败血杆菌(狗的犬咳，以及大多数哺乳动物的呼吸道疾病)。它的主要功能被认为是能够将其AC结构域转移到靶细胞的细胞膜上，并产生超生理学数量的cAMP。然而，我们现在已经证明，AC毒素形成跨膜孔的能力也与巨噬细胞的细胞毒性有关。此外，简单地将毒素与CR3结合，可以激活中性粒细胞(PMN)，而不依赖于cAMP的产生和毛孔的形成。CR3是一种白细胞整合素，可以作为AC毒素的受体。 在这个更新方案中，我们将发展这些最新的发现，并集中在AC毒素在建立呼吸道感染中对百日咳杆菌不可或缺的机制和靶向组织。我们将使用纯化的毒素或波氏杆菌(百日咳杆菌和败血杆菌)的体外方法来更多地了解毒素的组成、包装、分泌和传递(SA I)，以及毒素通过其三个功能(整合素受体占据、cAMP产生和毛孔形成)对两个主要候选靶组织--白细胞(SA II)和呼吸道上皮(SA III)的影响。最后，为了开始将AC毒素的作用与其他重要毒力因子的作用联系起来，我们将评估来自百日咳杆菌的其他毒素(百日咳毒素、气管细胞毒素)和粘附素(丝状血凝素、Pertactin)，以更好地了解它们对白细胞和呼吸道上皮(SA IV)可能的相互作用(增强、协同、拮抗或无作用)。其目的是应用关于AC毒素和其他波氏杆菌毒力因子的信息，更集中地研究AC毒素如何促进由该属引起的感染和疾病。

DESCRIPTION (provided by applicant): Adenylate cyclase (AC) toxin is a novel virulence factor for several species of the genus Bordetella, including B. pertussis (whooping cough, pertussis in humans), B. parapertussis (different strains causing diseases in animals and humans) and B. bronchiseptica (kennel cough in dogs, and respiratory diseases in most mammals). Its major function has been considered to be the ability to translocate its AC domain across the cytoplasmic membrane of target cells and produce supraphysiologic quantities of cAMP. We have now demonstrated, however, that the ability of AC toxin to form transmembrane pores is also relevant to cytotoxicity for macrophages. Furthermore, simply binding of the toxin to CR3, a leukocyte integrin that can act as a receptor for AC toxin, can activate human neutrophils (PMN), independent of its cAMP production and pore formation. In this renewal proposal, we will develop these recent discoveries and focus on the mechanisms and target tissues through which AC toxin is indispensable to B. pertussis in the establishment of infection in the respiratory tract. We will use in vitro approaches with either purified toxin or bordetella organisms (both B. pertussis and B. bronchiseptica) to learn more about toxin composition, packaging, secretion and delivery (SA I), the effects of the toxin on the two prime candidate target tissues, leukocytes (SA II) and the respiratory epithelium (SA III), through its three functions (occupancy of integrin receptor, cAMP production and pore formation). Finally, in order to begin to relate the actions of AC toxin to those of other important virulence factors, we will evaluate other toxins (pertussis toxin, tracheal cytotoxin) and adhesins (filamentous hemagglutinin, pertactin) from B. pertussis, both individually and in conjunction with AC toxin, to understand better the possible interactions (enhancement, synergy, antagonism or no effect) of their effects in leukocytes and the respiratory epithelium (SA IV). The objective is to apply the information on AC toxin and on other bordetellae virulence factors, to a more focused study of how AC toxin contributes to infections and diseases caused by this genus.