Novel Defensins of Human Epithelia

描述（由申请人提供）：抗菌肽的发现揭示了动物宿主防御的一个以前未被认识的组成部分。我们和其他人发现哺乳动物粘膜上皮细胞表达防御素肽，使它们具有参与粘膜宿主防御的能力。在人小肠中，防御素HD 5和HD 6在称为潘氏细胞的分泌上皮细胞中高度表达。我们小组和其他人的研究为潘氏细胞防御素肽在肠道中提供强大的关键防御功能的假设提供了强有力的支持。我们的研究还发现，HD 5作为修饰的前肽储存在潘氏细胞中。我们确定潘氏细胞胰蛋白酶是将proHD 5加工成存在于管腔中的活性抗微生物剂的蛋白酶。这些数据表明Paneth call胰蛋白酶的蛋白水解活性可能是调节肠道宿主防御的核心。 我们的假设是：1）HD 5 HD 6（和可能的HD 6）作为糖基化前肽储存在潘氏细胞中，提供了潜在抗生素活性和对宿主细胞的毒性降低的机制。在小肠中与HD 5一起具有（或协同的）抗微生物功能，和3）一旦被激活，潘氏细胞胰蛋白酶活性揭示潜在的抗菌剂并激活肠细胞上的蛋白酶激活受体2（PAR-2）。在三个具体目标中，我们将确定人proHD-5的组织形式的翻译后结构修饰（目标1）;建立HD-6的组织和腔形式的结构并确定它们的体外抗微生物活性（目标2）;表征潘氏细胞胰蛋白酶同工型，鉴定其关键活性，包括加工proHD-5和proHD-6，其他潜在抗菌剂的激活和PAR-2的激活，并确定潘氏细胞胰蛋白酶原激活的机制（目的3）。拟议的调查可能会提供新的治疗目标和方法，以打击肠道感染性和炎症性疾病的见解。

DESCRIPTION (provided by applicant): The discovery of antimicrobial peptides has unveiled a previously unrecognized component of animal host defense. We and others have discovered that mammalian mucosal epithelial cells express defensin peptides, providing them with the capacity to participate in mucosal host defense. In the human small intestine, defensins HD5 and HD6 are highly expressed in secretory epithelial cells called Paneth cells. Investigations by our group, and those of others, provide strong support for the hypothesis that Paneth cell defensin peptides provide a robust, critical defensive function in the intestine. Our studies have discovered also that HD5 is stored in Paneth cells as a modified propeptide. We identified that Paneth cell trypsin is the protease that processes proHD5 to the active antimicrobial present in the lumen. These data suggest that the proteolytic activity of Paneth call trypsin might be central to the regulation of enteric host defense. It is our hypothesis that 1) HD5 (and possibly HD6) is stored in Paneth cells as a glycosylated propeptide, providing a mechanism for both latent antibiotic activity and decreased toxicity to host cells, 2) HD6 has partially overlapping (or synergistic) antimicrobial functions with HD5 in the small intestine, and 3) once activated, Paneth cell trypsin activity unmasks latent antimicrobials and activates protease-activated receptor-2 (PAR-2) on enterocytes. In three specific aims, we will determine the posttranslational structural modifications of the tissue form of human proHD-5 (aim 1); establish the structure of tissue and luminal forms of HD-6 and determine their in vitro antimicrobial activity (aim 2); characterize Paneth cell trypsin isoforms, identify their key activities including processing of proHD-5 and -6, activation of other latent antimicrobials and activation of PAR-2, and identify the mechanism of Paneth cell protrypsin activation (aim 3). The proposed investigations may provide insights to novel therapeutic targets and approaches to combat infectious and inflammatory disease of the intestine.