Granulocyte-macrophage colony-stimulating factor (GM-CSF) has been studied for over forty years, and is among the most extensively researched cytokines. GM-CSF is involved in a variety of physiological processes ranging from wound healing and haematopoiesis to placental and fetal development, but it is best known for its role as a pro-inflammatory cytokine in immunity and inflammation. GM-CSF biology has been studied in various branches of immunology, such as autoimmunity, cancer, infectious diseases and vaccine development. In the immune system, GM-CSF has essential and non-redundant roles that make it an attractive therapeutic target. Several clinical trials targeting GM-CSF are underway, and initiation of additional trials is being considered. Both GM-CSF and its receptor knockout mice have been generated and have provided important insights into functioning of the immune system. However, transgenic mice that report GM-CSF expression do not exist, and Specific Aim 1 of this proposal is to generate such mice. We have designed genetic modifications in the GM-CSF reporter (Gr) mice that will readily enable subsequent generation of GM-CSF reporter/fate-reporter (Gr/fr) mice by crossing with existent transgenic mice, and this constitutes Specific Aim 2. The use of fate-reporter mice, which report gene expression even after expression ceases, has proved to be a valuable approach for studying questions that could not be adequately addressed otherwise. The principal conceptual innovation in the transgenic mice that we want to generate is the presence of two reporter genes, one that reports ongoing GM-CSF expression, the other reporting that GM-CSF has been expressed. This enables easy differentiation between current and past expression, which is not the case with fate-reporter mouse lines now being used. The Gr/fr mice will have normal GM-CSF expression, allowing studies of its biology in a physiological context.
In Specific Aim 3 we will address a relevant biological question that cannot be addressed without Gr/fr mice, exemplifying advances that can be made by use of these mice. Even though GM-CSF production is crucial for function of Th cells, a majority of them do not produce GM-CSF. It is not known whether subpopulations of Th cells have stable or transient phenotype with regard to GM-CSF expression, nor is it known which factors (a) determine Th cell expression of GM-CSF or (b) change its current GM-CSF- expressing status. We will test the hypothesis that subpopulations of Th cells have stable GM-CSF-expressing or -non-expressing phenotype. We expect that upon completion of this project we will have generated and comprehensively characterized Gr and Gr/fr mouse lines. We will also use Gr/fr mice to answer important biological questions, which will demonstrate their capacity to facilitate advances that would not be feasible otherwise.
Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a protein with a number of important functions in both normal and disease processes. We propose to generate genetically modified mice in which cells that produce GM-CSF will become fluorescent, allowing their easy identification and monitoring. Such mice will be a novel tool for advancing research, which may lead to development of new therapies.
