Granulocyte-macrophage colony-stimulating factor (GM-CSF) is an important cytokine which stimulates the growth, differentiation, and function of human myeloid cells. Although its biologic effects are normally attributed to a membrane-bound receptor, the recent discovery of a naturally occurring, soluble form of the GM-CSF receptor (sGMR-alpha) suggest that it too may play a role in regulating GM-CSF-mediated responses. Preliminary data from the PI suggest that sGMR-alpha may function differently from other soluble cytokine receptors in that it may associate with the signal transducing component of the GM-CSF receptor (betac) on the cell surface and do so in the absence of ligand. Thus sGMR-alpha may affect signal transduction in two ways; by binding GM-CSF in solution or by interfering with GM-CSF receptor aggregation. At present it is unclear whether sGMR- alpha in this context (or others) functions as a natural inhibitor or enhancer of GM-CSF signal transduction. The overall goal of this proposal is to define the structure, expression, and function of sGMR-alpha. In an effort to perform a comprehensive and detailed characterization, recombinant sGMR-alpha protein will be overexpressed and purified from eukaryotic cells. The recombinant protein will be analyzed for its ability to bind GM-CSF and associate with betac. These studies should provide insight into the biochemical mechanism of sGMR-alpha function and should extend our basic understanding of the structural dynamics of the GM-CSF receptor complex. In an effort to understand the physiological role of sGMR-alpha, a variety of cells and cell lines will surveyed for sGMR-alpha mRNA expression using reverse- transcriptase PCR (RT-PCR). The effects of different cytokines and inflammatory agents on sGMR-alpha expression will also be determined. These studies should indicate whether sGMR-alpha production in vivo is associated with promoting or inhibiting hematopoiesis and immune function. Expression studies will be verified in vitro by directly adding recombinant sGMR-alpha to a variety of cells and determining its effect on GM-CSF signal transduction. Cells to be analyzed include those engineered to exclusively express GMR-alpha or betac and those which naturally express the GM-CSF receptor such as proliferating myeloid cells, neutrophils, and nonhematopoietic tumor cell lines. Early, intermediate, and late events associated with GM-CSF signaling will be measured and should indicate the overall biologic importance of sGMR-alpha. Taken together, these studies should provide important information regarding the role of soluble cytokine receptors in vivo and should further our understanding of the molecular basis of hormone action. These studies may also set a precedent for utilizing sGMR-alpha as a therapeutic agent or a diagnostic indicator of disease.
| Kwon, E M; Raines, M A; Blenis, J et al. (2000) Granulocyte-macrophage colony-stimulating factor stimulation results in phosphorylation of cAMP response element-binding protein through activation of pp90RSK. Blood 95:2552-8 |
