TRANSCRIPTIONAL CONTROL OF INTERFERON-GAMMA IN T AND NK CELLS

性状（由申请方提供）：IFN-？，一种主要由NK细胞和Th 1CD 4和CD 8 T细胞产生的细胞因子，在对细胞内病原体感染的免疫应答中起中心作用。相反，对细胞外多细胞动物的保护依赖于Th 2 CD 4 T细胞和它们产生的细胞因子，包括IL-4、IL-13和IL-5。IFN-γ的表达而Th 2细胞因子主要在转录水平上调节。而调控IFN-？尽管近年来对表达方式进行了相当详细的阐述，但关于它们在何处以及如何起作用却知之甚少。与IFN-？相比，Th 2细胞因子IL-4、IL-13和IL-5在哺乳动物中聚集成单个~150 kb基因座，其调节现在在相当大的分子细节上被理解。在这个应用程序中，我们建议获得一个类似的知识水平有关IFN-？通过全面确定管理IFN-？表达和Ifng基因座的功能边界通过3个目的： 目的1：确定控制IFN-？通过对鼠Ifng基因座的全面染色质分析。假设：DNase超敏位点和区域丰富的转录有利的组蛋白修饰延伸约50 kb的上游和下游的小鼠IFNG基因，并确定管理IFN-？表情 目的2：定义的功能边界的Ifng基因座使用BAG转基因，以弥补缺陷的IFN-？敲除（GKO）小鼠。假设：含有鼠Ifng基因并延伸>= 50 kb 5'和3'的BAG转基因将重建适当的IFN-？表达，从而补充GKO小鼠中的免疫缺陷。 目的3：研究IFN-γ基因转录调控元件对IFN-γ表达的影响机制。表达和在这些位点起作用的关键转录因子。假设：T-bet、STAT 4、NFAT、NF-？B和AP-1不仅与Ifng启动子结合，还与其他上游和下游调控元件结合，从而影响IFN-？Ifng 5 ′ CNS 2和3和/或Ifng 3 ′ CNS 3有助于使Ifng基因座与周围基因的影响隔离。

DESCRIPTION (provided by applicant): IFN-?, a cytokine produced primarily by NK cells and by Th1 CD4 and CD8 T cells, plays a central role in the immune response to infection with intracellular pathogens. Conversely, protection against extracellular metazoans is dependent on Th2 CD4 T cells and the cytokines they produce, which include IL-4, IL-13 and IL-5. Expression of IFN-? and of Th2 cytokines is primarily regulated at the level of transcription. While transcription factors that govern IFN-? expression have in recent years been elucidated in considerable detail, very little is known regarding where and how they act. By contrast to IFN-?, regulation of the Th2 cytokines IL-4, IL-13 and IL-5, which are clustered into a single ~150 kb locus in mammals, is now understood in considerable molecular detail. In this application, we propose to gain a similar level of knowledge regarding IFN-? by comprehensively identifying the regulatory elements governing IFN-? expression and the functional boundaries of the Ifng locus through 3 Aims: Aim 1: Identify regulatory elements governing the expression of IFN-? through comprehensive chromatin profiling of the murine Ifng locus. Hypothesis: DNase hypersensitive sites and regions enriched in transcriptionally favorable histone modifications extend ~50 kb upstream and downstream of the murine Ifng gene and identify regulatory elements governing IFN-? expression. Aim 2: Define the functional boundaries of the Ifng locus using BAG transgenes to complement the defect in IFN-? knockout (GKO) mice. Hypothesis: BAG transgenes containing the murine Ifng gene and extending >= 50 kb 5' and 3' will reconstitute proper IFN-? expression and thereby complement the immunological defect in GKO mice. Aim 3: Identify the mechanisms bv which the transcriptional regulatory elements within the Ifng locus influence IFN-? expression and the key transcription factors that act at these sites. Hypotheses: T-bet, STAT4, NFAT, NF-?B and AP-1 bind not only to the Ifng promoter but to additional upstream and downstream regulatory elements, thereby affecting IFN-? expression; Ifng5'CNS 2 and 3 and/or Ifng3'CNS3 help to insulate the Ifng locus from the influence of surrounding genes.