Role of LITAF in Inflammatory Processes

LITAF 在炎症过程中的作用

• 批准号: 7540465
• 负责人: Salomon Amar
• 金额: $ 38.97万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-01-01 至 2011-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7540465
• 关键词: 3' Untranslated Regions; Address; Affect; Animal Model; Animals; Anti-Inflammatory Agents; Anti-inflammatory; Atherosclerosis; Binding; Binding Sites; Blood Vessels; Breeding; CCL2 gene; CCL8 gene; Cell Nucleus; Cells; Complex; Coupling; Crohn' s disease; Cytokine Gene; DNA Binding; DNA Sequence; DNA-Binding Proteins; DNA-Protein Interaction; Data; Defense Mechanisms; Development; Disease; Dose; Drops; Drug Design; Eicosanoids; Engineering; Escherichia coli; Evaluation; Family; Funding; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Goals; Human; Immune system; In Vitro; Infection; Inflammation; Inflammatory; Inflammatory Response; Injury; Insulin-Dependent Diabetes Mellitus; Interleukin-1; Interleukin-10; Interleukin-6; Knockout Mice; Leukocytes; Link; Lipopolysaccharides; Lipoproteins; Mediating; Mediator of activation protein; Modeling; Mus; Mutation; Names; Natural regeneration; Necrosis; Pathway interactions; Peptides; Periodontitis; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Play; Principal Investigator; Process; Production; Protein Binding; Protein Dephosphorylation; Protein Tyrosine Phosphatase; Proteins; Regulation; Reporter Genes; Research Personnel; Resolution; Rheumatoid Arthritis; Role; STAT6 gene; Sepsis; Septic Shock; Signal Pathway; Signal Transduction Pathway; Site; Syndrome; TLR2 gene; TLR4 gene; TNF gene; Testing; Tissues; Transcriptional Regulation; Transfection; Tumor Necrosis Factor-alpha; Tumor Necrosis Factors; Vascular Endothelial Growth Factors; Wound Healing; angiogenesis; cytokine; design; granulocyte; human TNF protein; in vivo; insight; macrophage; mitogen-activated protein kinase p38; mutant; novel; novel strategies; parallel processing; programs; promoter; protein aminoacid sequence; protein protein interaction; reactive oxygen intermediate; research study; response; success; tool; trafficking; transcription factor

Our long term goal is to pioneer new approaches that limit the harmful effects of inflammatory processes (such as those observed in periodontitis), while preserving the beneficial effects (e.g. tissue repair, resolution of infections). The studies we propose here build upon our recent results. Most notably, we discovered transcriptional regulators of inflammatory cytokines, including the pivotal cytokine tumor necrosis factor-a (TNF-a). TNF is tightly regulated; its overproduction can be lethal, as in septic shock syndrome. In previous studies we discovered a new transcription factor, Lipopolysaccharide-lnduced TNF-Alpha Factor (LITAF), which regulates TNF gene expression. More recently, we found another newly identified transcription factor STAT6B that binds to LITAF, and together this complex enhances the expression of a whole group of pro-inflammatory cytokines: GRO, IL-1a, RANTES, TNF-a, IFN-y, MCP-1, and MCP-2, VEGF as well as the anti-inflammatory cytokine IL-10. The Specific Aims of this proposal are designed to test the hypotheses that (1) LITAF stimulates the inflammatory response and together with STAT6B this response is largely amplified and (2) that STAT6B alone upregulates VEGF while together with LITAF, VEGF is inhibited. The proposed identification of the phosphorylation-dephosphorylation processes governing LITAF and STAT6B activity will be instrumental in understanding the cell trafficking of these molecules (Aim 1). The minimal and specific DNA sequence responsible for protein binding to LITAF on MCP1 promoter, and the role of LITAF-STAT6B (Aim 2), will allow identification of other promoters that may be targeted by these proteins. It will also permit evaluation of the role of LITAF and STAT6B in the regulation of their respective genes. To accomplish this, we will engineer specific mutations within the human MCP promoters, to reveal the exact DNA binding sequence controlled by LITAF and the role of STAT6B. Coupling the mutant promoters to reporter genes will allow us to assess whether these mutant promoters fail to be activated by LITAF and by the LITAF-STAT6B complex. Mutant LITAF and STAT6B proteins (muteins) will be designed to identify both the DNA-binding and trans-activation domains (Aim 2). and this data will provide critical insights for elucidating promoter interactions in other genes. A similar approach will be taken for STAT6B and VEGF to determine the minimal VEGF promoter sequence and STAT6B peptide involved in these DNA-protein interactions. The role of LITAF-STAT6B complex in inhibiting VEGF will be actively pursued in the same context (Aim 3). The success of LITAF knockout mice along with our proposal of generating STAT6B-deficient mice will build upon the results of Aims 1 and 2 and 3 to evaluate the hypothesis that the LITAF and STAT6B gene products, through regulation of cytokine activity, play an important role in the development of inflammatory diseases, including periodontitis (Aims 3). Aim 4 will expand our recent finding linking STAT6B to VEGF and angiogenesis. We intend to test how angiogenesis can be modulated by STAT6B and the LITAT-STAT6B complex. Our discovery of LITAF and the novel protein STAT6B, factors that help regulate cytokine transcription and angiogenesis, will serve as new tools to dissect the complex mechanisms that mediate cytokine expression in various inflammatory conditions, including periodontitis and vessel formation. Our goal is to develop pharmacological approaches aimed at modulating inflammation and angiogenesis.

我们的长期目标是开创限制炎症过程有害影响的新方法（例如那些 在牙周炎中观察到），同时保留有益效果（例如组织修复、感染解决）。研究 我们在此建议以我们最近的成果为基础。最值得注意的是，我们发现了炎症的转录调节因子 细胞因子，包括关键细胞因子肿瘤坏死因子-a (TNF-a)。 TNF 受到严格监管；其生产过剩 可能是致命的，如感染性休克综合征。在之前的研究中我们发现了一种新的转录因子， 脂多糖诱导的 TNF-α 因子 (LITAF)，调节 TNF 基因表达。最近，我们发现 另一种新发现的转录因子 STAT6B 与 LITAF 结合，该复合物共同增强了 整组促炎细胞因子的表达：GRO、IL-1a、RANTES、TNF-a、IFN-y、MCP-1 和 MCP-2， VEGF 以及抗炎细胞因子 IL-10。该提案的具体目标旨在测试 假设 (1) LITAF 刺激炎症反应，并且与 STAT6B 一起，这种反应 (2) STAT6B 单独上调 VEGF，而与 LITAF 一起则抑制 VEGF。 拟定的控制 LITAF 和 STAT6B 活性的磷酸化-去磷酸化过程的鉴定 将有助于理解这些分子的细胞运输（目标 1）。最小且特定的DNA 负责蛋白质与 MCP1 启动子上的 LITAF 结合的序列，以及 LITAF-STAT6B 的作用（目标 2），将允许 鉴定可能被这些蛋白质靶向的其他启动子。它还将允许评估 LITAF 的作用 和 STAT6B 对其各自基因的调节。为了实现这一目标，我们将在其中设计特定的突变 人类 MCP 启动子，以揭示 LITAF 控制的确切 DNA 结合序列以及 STAT6B 的作用。 将突变启动子与报告基因偶联将使我们能够评估这些突变启动子是否无法被 由 LITAF 和 LITAF-STAT6B 复合物激活。将设计突变的 LITAF 和 STAT6B 蛋白（突变蛋白） 识别 DNA 结合域和反式激活域（目标 2）。这些数据将为 阐明其他基因中启动子的相互作用。 STAT6B 和 VEGF 将采用类似的方法来确定 参与这些 DNA-蛋白质相互作用的最小 VEGF 启动子序列和 STAT6B 肽。的作用 LITAF-STAT6B 复合物抑制 VEGF 的作用将在相同的背景下积极研究（目标 3）。 LITAF 的成功 敲除小鼠以及我们生成 STAT6B 缺陷小鼠的建议将建立在目标 1 和 2 的结果的基础上 3 评估 LITAF 和 STAT6B 基因产物通过调节细胞因子活性的假设， 在包括牙周炎在内的炎症性疾病的发展中发挥着重要作用（目标 3）。目标4将扩大 我们最近的发现将 STAT6B 与 VEGF 和血管生成联系起来。我们打算测试如何调节血管生成 由 STAT6B 和 LITAT-STAT6B 复合物组成。我们发现了 LITAF 和新型蛋白质 STAT6B，这些因素 帮助调节细胞因子转录和血管生成，将作为剖析该复合物的新工具 在各种炎症条件下介导细胞因子表达的机制，包括牙周炎和 血管形成。我们的目标是开发旨在调节炎症和 血管生成。