Translational regulation of tissue resident macrophages by GCN2

总结 mRNA翻译或蛋白质合成是一个基本的细胞过程，在几种情况下可能会失调。 人类疾病。巨噬细胞是存在于大多数组织中的异质群体， 组织特异性功能。mRNA翻译控制在巨噬细胞及其组织中的作用 具体功能还不清楚。拟议研究的主要目标是了解如何 mRNA翻译的失调控制应激期间组织驻留的巨噬细胞功能。具体 本研究的目的是确定GCN 2（一般控制非去阻遏蛋白2）依赖性翻译 控制巨噬细胞影响巨噬细胞的功能，在红细胞的生产和清除，并揭示 基因介导这种效应。GCN 2是属于信号网络的丝氨酸/苏氨酸蛋白激酶 通过翻译调节因子eIF 2（真核翻译）协调细胞对营养胁迫的反应 起始因子2）。GCN 2传感氨基酸水平并响应氨基酸水平磷酸化eIF 2 缺陷p-eIF 2抑制整体mRNA的翻译，但矛盾的是，它刺激了一个子集的翻译。 关键的应激反应基因，如ATF 4（激活转录因子4）。应激反应上调 响应GCN 2/eIF 2信号的基因激活了一个转录程序，帮助细胞克服 不利的条件或经历凋亡。GCN 2功能以前曾与重要的 生理和病理状况，如记忆形成、癌症和炎性疾病。 然而，GCN 2在调节组织驻留的巨噬细胞中的作用及其在RBC产生中的功能， 清除率尚未确定。我们目前的模型表明，GCN 2控制红细胞的产生， 以及通过调节巨噬细胞中的mRNA翻译在应激期间清除。所以我们提出 为了达到我们的目标，我们将首先通过以下方法确定GCN 2在RBC清除中的重要性： 巨噬细胞（目标1），并定义GCN 2影响这一过程的分子机制。接下来我们 将阐明GCN 2如何控制红细胞成熟和巨噬细胞的生产（目的2）。最后我们将 通过GCN 2检测巨噬细胞对机械力的感受（目的3）。为了实现这些目标，我们将 使用缺乏GCN 2或在巨噬细胞中携带磷酸化抗性eIF 2的转基因小鼠， 在全基因组水平上研究mRNA翻译的技术。我们的小鼠模型以及体内和体外 实验将严格评估巨噬细胞在GCN 2依赖性肿瘤发生中的中心作用。 压力下的缺陷。我们的全基因组方法和选定靶点的体外功能分析将 发现在巨噬细胞中起重要作用的GCN 2下游的新的免疫调节基因 在应激期间调节RBC产生。

SUMMARY mRNA translation, or protein synthesis, is a fundamental cellular process that can be dysregulated in several human diseases. Macrophages are heterogeneous populations that are present in most tissues and adopt tissue specific functions. The role of mRNA translational control in macrophages and in regards to their tissue specific functions is not well understood. The broad goal of the proposed studies is to understand how dysregulation of mRNA translation controls tissue-resident macrophage function during stress. The specific goals of this study are to identify how GCN2 (general control nonderepressible 2)-dependent translational control in macrophages affects macrophage function in RBC production and clearance and to uncover the genes mediating this effect. The GCN2 is a serine/threonine-protein kinase that belongs to a signaling network that coordinates cellular response to nutrient stress through translational regulator eIF2 (Eukaryotic translation initiation factor 2). GCN2 senses amino acid levels and phosphorylates eIF2 in response to amino acid deficiency. p-eIF2 inhibits global mRNA translation but paradoxically stimulates the translation of a subset of key stress-response genes such as ATF4 (Activating Transcription Factor 4). Upregulation of stress-response genes in response to GCN2/eIF2 signaling activates a transcription program that helps the cells to overcome unfavorable conditions or undergo apoptosis. GCN2 function has been previously linked to important physiological and pathological conditions such as memory formation, cancer and inflammatory diseases. However, the role of GCN2 in regulating tissue-resident macrophages and their functions in RBC production and clearance has not been characterized. Our current model suggests that GCN2 controls RBC production and clearance during stress through regulation of mRNA translation in macrophages. Therefore, we propose the following aims to achieve our goals: First we will determine the importance of GCN2 in RBC clearance by macrophages (Aim 1) and define molecular mechanisms through which GCN2 impact this process. Next, we will elucidate how GCN2 controls RBC maturation and production by macrophages (Aim 2). Finally, we will examine how mechanical force sense by macrophage through GCN2 (Aim 3). To achieve these goals we will use transgenic mice lacking GCN2 or carrying phospho-resistant eIF2 in macrophages and state-of-art technology to study mRNA translation at genome-wide level. Our mouse models and in vivo and in vitro experiments will rigorously assess the central role of macrophages in development of GCN2-dependent defects during stress. Our genome-wide approach and in vitro functional analysis of selected targets will discover novel translationally regulated genes downstream of GCN2 that play important roles in macrophage regulation of RBC production during stress.