Translational regulation of tissue resident macrophages by GCN2

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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.