ISGF3 TRANSCRIPTION FACTOR FAMILY IN CYTOKINE SIGNALING

细胞因子信号转导中的 ISGF3 转录因子家族

• 批准号: 2855980
• 负责人: David E Levy
• 金额: $ 37.15万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-01-01 至 1999-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2855980
• 关键词: DNA binding protein; JAK kinase; biological signal transduction; cytokine; embryonic stem cell; epidermal growth factor; flow cytometry; gene expression; gene targeting; genetically modified animals; growth factor receptors; immunoprecipitation; interferons; laboratory mouse; light microscopy; molecular cloning; northern blottings; platelet derived growth factor; polymerase chain reaction; protein signal sequence; protein structure function; receptor binding; receptor coupling; restriction fragment length polymorphism; site directed mutagenesis; tissue /cell culture; transcription factor; western blottings

The mechanisms of signal transduction from cytokine and growth factor receptors are important for understanding the physiological roles of these important mediators of cell growth, differentiation, and response to pathogens. We have been defining the biochemistry of signal transduction for interferons (IFNs) and have extended the paradigm discovered in this system to the epidermal and platelet-derived growth factors (EGF & PDGF). The proposed research will define, clone and characterize novel transcription factors regulated by these cytokines; determine the biochemical mechanisms involved in coupling cell surface receptors to activation of transcription; identify and isolate target genes activated by cytokines functioning through this pathway; and create mutant cell lines and mouse strains specifically deficient in this signaling pathway by gene targeting through homologous recombination. Understanding the biochemical mechanisms of growth factor and cytokine signaling and their target genes will allow definition of steps deregulated in pathological states such as cancer where over activity of growth factors or under activity of growth inhibitory cytokines leads to abnormal cell growth. Creation of mutant cell lines and animals with specific lesions in cytokine pathways will define the physiological consequences of cytokine functioning under normal and abnormal conditions and will provide new models for disease. New members of the ISGF3 transcription factor family will be cloned and analyzed for structure-function relationships, for their ability to form multimers with novel DNA-binding specificity, for their ability to be activated by different cytokines, and for their differential, tissue- specific patterns of expression. The biochemistry of signal coupling allowing cytokine and growth factor receptors to activate transcription factors will be defined by determining the linkage between receptors, the Jak family kinases, and the transcription factor substrates. Structure- function studies of the EGF and PDGF receptor and Jak1 kinase will define physical interactions linking these proteins and the mechanisms of ligand- induced activation. Similarly, studies of the p9l and p93 proteins of ISGF3 will define how they are specifically recognized by growth factor and/or IFN receptors. Target genes activated by growth factors will be identified by arbitrarily-primed PCR fingerprinting of RNA from cells treated under different conditions. Expression patterns from wildtype and mutant EGF and PDGF receptors will define genes regulated via the ISGF3 pathway. Altered embryonic stem cells with disruption of the ISGF3 p9l gene by homologous recombination will be used to create homozygous mutant animals. Homozygous null cells will be analyzed for their capacity to differentiate in vitro and for ability to colonize specific tissues in chimeric mice, with particular attention to hematopoiesis.

细胞因子和生长因子的信号转导机制 受体对于理解这些受体的生理作用是重要的。 细胞生长、分化和应答的重要介质 病原体我们已经定义了信号传导的生物化学 对于干扰素（IFN），并已扩展了本研究中发现的范式， 表皮和血小板衍生生长因子（EGF和PDGF）。 拟议的研究将定义，克隆和表征新的 由这些细胞因子调节的转录因子;确定 参与偶联细胞表面受体的生化机制， 转录激活;鉴定和分离激活的靶基因 细胞因子通过这一途径发挥作用;并创造突变细胞 细胞系和小鼠品系特异性地缺乏该信号通路 通过同源重组进行基因打靶。了解 生长因子和细胞因子信号转导的生化机制及其 靶基因将允许定义病理学中失调的步骤， 如癌症，其中生长因子的活性过高或 生长抑制性细胞因子的活性导致异常细胞生长。 产生突变细胞系和具有特定病变的动物， 细胞因子途径将定义细胞因子的生理后果 在正常和异常情况下运行，并将提供新的 疾病的模型。 ISGF 3转录因子家族的新成员将被克隆， 分析了结构-功能关系，分析了它们形成 具有新的DNA结合特异性的多聚体，因为它们能够 由不同的细胞因子激活，对于它们的差异，组织- 具体的表达方式。信号耦合的生物化学 允许细胞因子和生长因子受体激活转录 因子将通过确定受体之间的联系来定义， Jak家族激酶和转录因子底物。结构- EGF和PDGF受体和Jak 1激酶的功能研究将定义 连接这些蛋白质和配体机制的物理相互作用- 诱导激活。类似地，对p9 l和p93蛋白的研究表明， ISGF 3将定义它们如何被生长因子特异性识别 和/或IFN受体。被生长因子激活的靶基因将被 通过来自细胞的RNA的任意引物PCR指纹图谱鉴定 在不同的条件下处理。来自野生型的表达模式和 突变的EGF和PDGF受体将定义通过ISGF 3调节的基因 通路改变的胚胎干细胞与ISGF 3 p9 l的破坏 基因通过同源重组将被用于创建纯合突变体 动物将分析纯合无效细胞的能力， 在体外分化和定殖特定组织的能力， 嵌合小鼠，特别关注造血。