Dendritic cells (DCs) provide a critical connection between the innate and adaptive immune responses by receiving signals from pathogens via pattern recognition receptors and transmitting signals that activate na?ve T cells, NK and B cells. Several DC subsets exist, which display important differences in function. Plasmacytoid dendritic cell precursors (pDCs) are professional type I interferon-producing cells, while conventional or myeloid dendritic cells (mDCs) demonstrate potent antigen presentation function. The pathways that control development of these DC subsets are unknown. Once elucidated, this information may provide methods to control or redirect immune responses during infection, disease or for therapeutic applications. The goal of this project is to understand the molecular regulation of pDC and mDC development by cytokines. Maturation of pDCs is critically dependent on Flt3 ligand (Flt3L) and its downstream signaling protein STAT3. Flt3L-dependent pDC development is severely inhibited by granulocyte-macrophage colony- stimulating factor (GM-CSF), which promotes mDC formation at the expense of pDCs. The suppressive activity of GM-CSF on pDC development operates at the progenitor pDC (pro-pDC) stage and requires the transcription factor STAT5. These findings led to the hypothesis that GM-CSF-activated STAT5 stimulates a transcriptional program that represses Flt3-dependent STAT3 signaling and/or expression of critical lineage- specification factors, thus abrogating pDC development.
Two aims are proposed to test this hypothesis.
In Aim 1, the role of GM-CSF and STAT5 in modulating Flt3 function in pDCs and pro-pDCs will be determined. Cellular responses to GM-CSF will be examined by measuring the survival, growth and absolute production of pDCs and mDCs from wild-type or STAT5-deficient pro-pDCs in Flt3L cultures containing or lacking GM-CSF. Molecular mechanisms of GM-CSF will be evaluated by examining Flt3 and GM-CSF receptor expression, Flt3/STAT3 signal transduction, and expression of SOCS family negative regulators in wild type and STAT5- deficient cells.
In Aim 2, the effect of GM-CSF signaling and STAT5 activity on the pDC and mDC transcriptional regulatory networks will be examined. Candidate DC and STAT target gene expression will be measured during development of wild type, STAT5- and STAT3-deficient pDCs and mDCs. This project will reveal regulatory pathways of DC development by cytokines. ? ? ?
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