Defining Protective Responses in Hematopoietic Cells Mediated by STAT3 Anti-Inflammatory Activity

项目总结/摘要 造血系统是炎症反应的基础。造血干细胞和 存在于成人骨髓中的多能祖细胞（统称为HSPC）产生炎性细胞毒性。 骨髓和淋巴细胞群。HSPC还通过以下方式对病原体相关的炎性刺激作出反应： 调节抵抗感染所需的细胞输出。不幸的是，HSPC活性在慢性 炎性条件。这些诱导HSPC失调，丧失长期干细胞重建功能， DNA损伤和骨髓造血异常在人类中，慢性炎症与骨骼有关 骨髓衰竭、骨髓增生异常综合征和急性髓性白血病。一个重要的知识缺口是如何 保护造血系统免受炎症的有害影响。通过研究这个问题， 我们发现了一个关键的作用，为精氨酸激活转录调节STAT 3;具体来说，我们的研究结果 提示STAT 3抗炎活性对于保存HSPC和谱系平衡是必需 造血我们以前发现STAT 3的抗炎活性是由转录调控因子介导的。 抑制Ube 2n，编码Ubc 13，一种激活TRAF 6和NF-κ B的E2泛素结合酶， Toll样受体（TLR）下游的κB/MAPK信号传导（例如，TLR2、TLR4）。我们发现了造血- 限制性Stat 3缺乏导致骨髓衰竭、HSPC损失、骨髓偏斜造血和 增加的祖细胞促炎信号传导。引人注目的是，同时从Stat 3缺陷型中去除Ube 2n， 造血细胞使造血活性，这表明需要STAT 3抑制Ubc 13， 保持功能性HSPC。因此，我们假设STAT 3在保护HSPCs免受 炎症诱导的损伤通过调节Ubc 13。为了验证这一点（目标1），我们将研究 STAT 3在炎症中保护造血功能。使用混合骨髓嵌合体，我们将 确定STAT 3和Ubc 13在炎症造血反应中的作用;我们将研究STAT 3- 激活骨髓中的细胞因子和生产者群体;我们将测试STAT 3的需求 使用表达转录抑制因子的转基因菌株， 缺陷型STAT 3突变体。在目标2中，我们将描述STAT 3保护HSPCs的分子途径 炎症引起的损伤我们将确定STAT 3和Ubc 13在保护HSPC免受 炎症过程中DNA损伤和静止期丧失，我们将确定STAT 3和Ubc 13基因组， HSPC在稳态和炎症中的广泛转录应答。我们预计该项目将 提供了前所未有的洞察内在HSPC保护机制，基本信息，将 推动该领域向前发展，以提高对炎症期间免疫系统调节的理解。

PROJECT SUMMARY/ABSTRACT The hematopoietic system is the foundation of the inflammatory response. Hematopoietic stem cells and multipotent progenitors (collectively termed HSPCs) residing in adult bone marrow generate inflammatory myeloid and lymphoid populations. HSPCs also respond to pathogen-associated inflammatory stimuli by adjusting the output of cells required to fight infection. HSPC activity is unfortunately co-opted in chronic inflammatory conditions. These induce HSPC deregulation, loss of long-term stem cell repopulating function, DNA damage, and myeloid-skewed hematopoiesis. In humans, chronic inflammation is linked with bone marrow failure, myelodysplastic syndrome, and acute myeloid leukemia. An important gap in knowledge is how the hematopoietic system is protected from the harmful effects of inflammation. By investigating this question, we discovered a key role for the cytokine-activated transcriptional regulator STAT3; specifically, our results suggest STAT3 anti-inflammatory activity is necessary to preserve HSPCs and lineage-balanced hematopoiesis. We showed previously that STAT3 anti-inflammatory activity is mediated by transcriptional repression of Ube2n, encoding Ubc13, an E2 ubiquitin-conjugating enzyme that activates TRAF6 and NF- κB/MAPK signaling downstream of Toll-like receptors (TLRs) (e.g., TLR2, TLR4). We found hematopoietic- restricted Stat3-deficiency leads to bone marrow failure, HSPC loss, myeloid-skewed hematopoiesis and increased progenitor pro-inflammatory signaling. Strikingly, concomitant removal of Ube2n from Stat3-deficient hematopoietic cells enables hematopoietic activity, suggesting STAT3 restraint of Ubc13 is required to maintain functional HSPCs. Thus, we hypothesize STAT3 has a central role in protecting HSPCs from inflammation-induced damage via modulation of Ubc13. To test this (Aim 1), we will investigate the role for STAT3 in protecting hematopoietic function in inflammation. Using mixed bone marrow chimeras, we will determine roles for STAT3 and Ubc13 in hematopoietic responses to inflammation; we will investigate STAT3- activating cytokines and producer populations in bone marrow; and we will test the requirement for STAT3 transcriptional activity in HSPCs during inflammation using a transgenic strain expressing a transcriptionally defective STAT3 mutant. In Aim 2, we will delineate molecular pathways by which STAT3 protects HSPCs from inflammation-induced damage. We will determine roles for STAT3 and Ubc13 in protecting HSPCs from DNA damage and loss of quiescence during inflammation, and we will determine STAT3- and Ubc13- genome- wide transcriptional responses in HSPCs in homeostasis and inflammation. We anticipate this project will provide unprecedented insight into intrinsic HSPCs protective mechanisms, fundamental information that will move the field forward to an improved understanding of immune system regulation during inflammation.