Role and Regulation of a Molecular Mimic in Histoplasma Pathogenesis

描述（由申请人提供）：荚膜组织浆体是一种经典的“二态”真菌病原体，经历温度诱导从生长在土壤中的霉菌形式转变为在肺巨噬细胞中建立感染的酵母菌形式。所有的二态真菌在免疫功能低下的患者中引起最严重的疾病，但它们是原发病原体，可以在没有任何明显免疫缺陷的宿主中引起严重问题。第一个被遗传学证实的荚膜芽胞杆菌毒力因子是CBP，这是一种酵母期特异性分泌蛋白，对体外和体内巨噬细胞的增殖都很重要。解决CBP的三维结构揭示了其与皂苷B的惊人相似性，皂苷B是一种参与膜加工和抗原呈递的哺乳动物蛋白。尽管结构同源性强烈提示CBP是第一个被发现的庞大而多样的皂苷样蛋白家族的真菌成员，但其在组织胞浆菌病中的确切机制作用仍未得到证实。该资助申请旨在评估CBP是否确实作为皂苷起作用，并了解CBP1阶段特异性调控的基础。定义CBP的皂苷样功能。本Aim的研究将首先测试CBP在结合脂质方面是否像皂苷一样起作用，无论是真菌还是哺乳动物，并将包括一个

DESCRIPTION (provided by applicant): Histoplasma capsulatum is one of the classic "dimorphic" fungal pathogens, undergoing a temperature-induced transition from a mold form that grows in soil to a yeast form that establishes infection in lung macrophages. All of the dimorphic fungi cause the most severe disease in immunocompromised patients, but they are primary pathogens that can cause serious problems in hosts lacking any demonstrable immune defect. The first genetically proven virulence factor of H. capsulatum was CBP, a yeast phase-specific secreted protein that is important for proliferation within macrophages in vitro and in vivo. Solving the three-dimensional structure of CBP revealed a surprising similarity to saposin B, a mammalian protein involved in membrane processing and antigen presentation. Although the structural homology is strongly suggestive that CBP is the first identified fungal member of the large and diverse saposin-like protein family, its precise mechanistic role in histoplasmosis remains unproven. This grant application is designed to evaluate whether CBP indeed functions as a saposin and to understand the basis for CBP1 phase-specific regulation: Specific Aim 1. Define the saposin-like function of CBP. The studies in this Aim will first test whether CBP functions like a saposin in terms of binding lipids, either fungal or mammalian, and will include a structure-function analysis to determine which residues of CBP are involved. The potential role of CBP in CD1 antigen presentation, analogous to the function of saposin B, will also be explored. Specific Aim 2. Identify the genes involved in CBP1 transcriptional regulation. Insertional mutagenesis will be used to generate a library of mutants that will be screened for those that no longer regulate CBP1 normally. The most promising candidate regulatory genes will subsequently be used to identify coordinately regulated genes, potentially unraveling a network of genes related to CBP function or to other roles in pathogenesis.