ABSTRACT It has been appreciated recently that tissue-resident macrophages (TRMs) contribute to tissue homeostasis. For example, in the brain, microglia have been proposed to process extracellular proteins and to restrict the formation of plaque formed from proteins such as -amyloid and Tau proteins. Such tissue-resident macro- phages were once thought to be derived from monocytes that arise in definitive hematopoiesis in the adult bone marrow. However, we now understand that TRMs are derived during fetal life and arise from progenitors in the yolk sac and fetal liver. In some tissues, these embryonically derived macrophage populations persist throughout adult life and are not efficiently replaced by circulating monocytes arising from adult bone marrow and definitive hematopoiesis. One impediment to studying the functions of such embryonically derived macro- phages is the relative paucity of information regarding their development, specifically the transcriptional pro- grams that guide their development. The lack of such information prevents the design of model systems where these cells can be prevented from developing, which would allow their in vivo functions to be defined in a defin- itive way. The current application is aimed at defining the basis for the embryonic development of macrophag- es. It is based on preliminary data that has: 1) identified the Zeb2 enhancer (at -165kb) used in definitive hematopoiesis supporting monocyte/macrophage development from the adult bone marrow; 2) showed that this enhancer is not required for Zeb2 expression in embryonic macrophage development, and 3) identified two other Zeb2 enhancers that are selectively active in the embryonic yolk sac and fetal liver progenitors. This application will (Aim 1) test these embryonically active Zeb2 enhancers for their role in supporting embry- onic macrophage development, and (Aim 2) determine the transcriptional basis for their embryonic activity. Aim 1 may directly produce models of embryonic macrophage deficiency, which would immediately benefit re- search in other areas related to immune cell support of tissue homeostasis. Beyond this, the basic information on the similarities or differences between primitive and adult hematopoiesis could have further uses, such as in adding to ways in which fetal gene expression can be controlled to compensate for innate or acquired defects in adult gene expression, such as in -thalassemia.