STAT1-DEPENDENT TRANSCRIPTION IN AIRWAY EPITHELIAL CELLS

描述 (改编自申请人的摘要)申请人建议航空公司 上皮细胞为调控提供关键的生化信号 免疫和炎症。为了支持这种可能性，他们发现 上皮细胞表达一套免疫反应基因的功能 (ICAM-1、IRF-1、TAP1和RANTES)在干扰素-γ的控制下 (干扰素、γ)信号转导途径。例如，上皮细胞 细胞间黏附分子-1(ICAM-1)的表达介导白细胞 呼吸道内的贩运(粘连、移行和滞留) 它通过作为白细胞B2整合素的受体作用于上皮细胞。在那 背景下，他们已经获得了ICAM-1介导上皮免疫的证据 细胞间的相互作用可允许产生“适当的”免疫反应(例如， 呼吸道病毒感染)或“不适当的”反应(例如，呼吸道 哮喘的炎症)。这个应用程序基于观察到的 ICAM-1在呼吸道上皮细胞上的表达依赖于特定的 干扰素-γ驱动的信号转导通路依次依赖于 激活的转录因子STAT1与ICAM-1基因的结合 启动子区域。除了这种DNA/蛋白质相互作用外，STAT1还 似乎与转录辅活化子p300和 转录因子Sp1充分激活基因转录。的目的是 此应用程序用于确定STATL的结构特征，以允许 关于与p3OO和Sp1的相互作用以及这些相互作用在 激活ICAM-1及相关免疫反应基因的转录。这 然后，可以使用信息来修改ICAM-1基因的激活和呼吸道 阻断增强子通路和增强的炎症反应 抑制子途径(例如，STATL显性-负性突变和腺病毒 EIA蛋白表达)。 因此，具体目标是：一、界定结构基础和 STAT1/p3OO相互作用在ICAM-1基因激活中的作用他们 将使用他们的原代培养的人类呼吸道上皮细胞模型来定义 STAT1与转录辅活化子p3OO的相互作用。 STAT1和p3OO的结构和功能研究将于#年完成 定义蛋白质/蛋白质相互作用和基因转录的实验 用于野生型和突变形式的他汀类和p3OO。二、界定 ICAM-1基因中STATL/SPL-相互作用的结构基础和功能作用 激活。他们将利用相同的细胞模型来表征 转录协同所需的结构域 STATL和SPL。STAT1和Sp1的结构和功能研究将 通过蛋白质/蛋白质、蛋白质/DNA和基因转录完成 野生型和突变型他汀类药物的实验。(末尾 摘要)

DESCRIPTION (Adapted from applicants' abstract) The applicants propose that airway epithelial cells provide critical biochemical signals for regulating immunity and inflammation. In support of this possibility, they have found that epithelial cells express a functional set of immune-response genes (ICAM-1, IRF-1, TAP1, and RANTES) under the control of an interferon-gamma (IFN-,gamma) signal transduction pathway. For example, epithelial cell expression of intercellular adhesion molecule-1 (ICAM- 1) mediates leukocyte trafficking (adhesion, transmigration, and retention) in the airway epithelium by serving as a receptor for leukocyte B2-integrins. In that context, they have obtained evidence that ICAM-1 mediates epithelial-immune cell interactions that may allow for "appropriate" immune responses (e.g., to a respiratory virus infection) or "inappropriate" responses (e.g., airway inflammation in asthma ). This application is based on the observation that ICAM-1 expression on airway epithelial cells depends on a specific IFN-gamma-driven signal transduction pathway that relies in turn on the binding of the activated transcription factor Statl to the ICAM-1 gene promoter region. In addition to this DNA/protein interaction, Statl also appears to interact with the transcriptional coactivator p3OO and the transcription factor Sp1 to fully activate gene transcription. The aim of this application is to determine the structural features of Statl that allow for interaction with p3OO and Sp1 and the role of these interactions in activating transcription for ICAM-1 and related immune-response genes. This information may then be used to modify ICAM-1 gene activation and the airway inflammatory response through blockade of enhancer-pathways and potentiation of repressor-pathways (e.g., Statl dominant-negative mutation and adenoviral EIA protein expression). Accordingly, The specific aims are to: I. Define the structural basis and functional role of Statl/p3OO interaction in ICAM-1 gene activation. They will use their primary culture human airway epithelial cell model to define the interaction between Statl and the transcriptional coactivator p3OO. Structure-function studies of Statl and p3OO will be accomplished in experiments that define protein/protein interactions and gene transcription for wild-type and mutated forms of Statl and p3OO. II. Define the structural basis and functional role of Statl/Spl-interaction in ICAM-1 gene activation. They will take advantage of the same cell model to characterize the structural domains required for the transcriptional synergy between Statl and Spl. Structure-function studies of Stat1 and Sp1 will be accomplished using protein/protein, protein/DNA, and gene transcription experiments for wild-type and mutated forms of Statl and Spl. (End of Abstract)