Cytokine Regulation of Dendritic Cell Development

描述(申请人提供)：树突状细胞(DC)通过模式识别受体接收来自病原体的信号，并传递激活NAVE T细胞、NK细胞和B细胞的信号，从而在先天免疫反应和获得性免疫反应之间提供关键连接。存在几个DC亚集，它们在功能上显示出重要的差异。浆细胞样树突状细胞前体(PDC)是专职的I型干扰素产生细胞，而常规或髓系树突状细胞(MDCs)具有强大的抗原提呈功能。控制这些DC亚群发展的途径尚不清楚。一旦被阐明，这些信息可能提供在感染、疾病或治疗应用期间控制或重定向免疫反应的方法。本项目的目标是了解细胞因子对PDC和MDC发育的分子调控。PDC的成熟依赖于Flt3配体(Flt3L)及其下游信号蛋白STAT3。粒细胞-巨噬细胞集落刺激因子(GM-CSF)以牺牲pDC为代价促进mDC的形成，严重抑制依赖Flt3L的pDC的形成。GM-CSF对PDC发育的抑制作用在前体PDC(Pro-PDC)阶段起作用，需要转录因子STAT5。这些发现导致了一种假设，即GM-CSF激活的STAT5刺激转录程序，抑制依赖于Flt3的STAT3信号和/或关键谱系指定因子的表达，从而消除PDC的发育。为了检验这一假设，本文提出了两个目标。在目标1中，将确定GM-CSF和STAT5在pDC和Pro-pDC中调节Flt3功能的作用。在含有或不含GM-CSF的Flt3L培养中，将通过测量野生型或STAT5缺陷的pDC和MDCs的存活、生长和绝对产量来检测细胞对GM-CSF的反应。GM-CSF的分子机制将通过检测Flt3和GM-CSF受体的表达，Flt3/STAT3信号转导，以及SOCS家族负调控因子在野生型和STAT5缺陷细胞中的表达来评估。在目标2中，将检测GM-CSF信号和STAT5活性对PDC和MDC转录调控网络的影响。在野生型、STAT5和STAT3缺失的pDC和MDCS的发育过程中，将测量候选DC和STAT靶基因的表达。该项目将揭示细胞因子对DC发育的调控途径。

DESCRIPTION (provided by applicant): Dendritic cells (DCs) provide a critical connection between the innate and adaptive immune responses by receiving signals from pathogens via pattern recognition receptors and transmitting signals that activate na¿ve T cells, NK and B cells. Several DC subsets exist, which display important differences in function. Plasmacytoid dendritic cell precursors (pDCs) are professional type I interferon-producing cells, while conventional or myeloid dendritic cells (mDCs) demonstrate potent antigen presentation function. The pathways that control development of these DC subsets are unknown. Once elucidated, this information may provide methods to control or redirect immune responses during infection, disease or for therapeutic applications. The goal of this project is to understand the molecular regulation of pDC and mDC development by cytokines. Maturation of pDCs is critically dependent on Flt3 ligand (Flt3L) and its downstream signaling protein STAT3. Flt3L-dependent pDC development is severely inhibited by granulocyte-macrophage colony- stimulating factor (GM-CSF), which promotes mDC formation at the expense of pDCs. The suppressive activity of GM-CSF on pDC development operates at the progenitor pDC (pro-pDC) stage and requires the transcription factor STAT5. These findings led to the hypothesis that GM-CSF-activated STAT5 stimulates a transcriptional program that represses Flt3-dependent STAT3 signaling and/or expression of critical lineage- specification factors, thus abrogating pDC development. Two aims are proposed to test this hypothesis. In Aim 1, the role of GM-CSF and STAT5 in modulating Flt3 function in pDCs and pro-pDCs will be determined. Cellular responses to GM-CSF will be examined by measuring the survival, growth and absolute production of pDCs and mDCs from wild-type or STAT5-deficient pro-pDCs in Flt3L cultures containing or lacking GM-CSF. Molecular mechanisms of GM-CSF will be evaluated by examining Flt3 and GM-CSF receptor expression, Flt3/STAT3 signal transduction, and expression of SOCS family negative regulators in wild type and STAT5- deficient cells. In Aim 2, the effect of GM-CSF signaling and STAT5 activity on the pDC and mDC transcriptional regulatory networks will be examined. Candidate DC and STAT target gene expression will be measured during development of wild type, STAT5- and STAT3-deficient pDCs and mDCs. This project will reveal regulatory pathways of DC development by cytokines.