Cytokine Regulation of Dendritic Cell Development

描述（由申请人提供）：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初始T细胞、NK细胞和B细胞的信号，在先天性和适应性免疫应答之间提供关键联系。存在几个DC子集，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）是专业的I型干扰素产生细胞，而常规或髓样树突状细胞（mDC）表现出有效的抗原呈递功能。控制这些DC亚群发育的途径尚不清楚。一旦阐明，该信息可以提供在感染、疾病期间控制或重定向免疫应答或用于治疗应用的方法。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的成熟严重依赖于Flt 3配体（Flt 3L）及其下游信号传导蛋白STAT 3。Flt 3L依赖性pDC发育受到粒细胞-巨噬细胞集落刺激因子（GM-CSF）的严重抑制，粒细胞-巨噬细胞集落刺激因子（GM-CSF）以pDC为代价促进mDC形成。GM-CSF对pDC发育的抑制活性在祖细胞pDC（pro-pDC）阶段起作用，并且需要转录因子STAT 5。这些发现导致以下假设：GM-CSF激活的STAT 5刺激抑制Flt 3依赖性STAT 3信号传导和/或关键谱系特化因子表达的转录程序，从而消除pDC发育。提出了两个目标来检验这一假设。在目的1中，将确定GM-CSF和STAT 5在pDC和pro-pDC中调节Flt 3功能的作用。将通过测量在含有或缺乏GM-CSF的Flt 3L培养物中来自野生型或STAT 5缺陷型pro-pDC的pDC和mDC的存活、生长和绝对产生来检查对GM-CSF的细胞应答。将通过检查野生型和STAT 5缺陷型细胞中Flt 3和GM-CSF受体表达、Flt 3/STAT 3信号转导和SOCS家族负调节因子的表达来评价GM-CSF的分子机制。在目的2中，将检查GM-CSF信号传导和STAT 5活性对pDC和mDC转录调控网络的影响。将在野生型、STAT 5和STAT 3缺陷型pDC和mDC的开发期间测量候选DC和STAT靶基因表达。本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.