The Role of GM-CSF Signaling and CREB in Myeloid Proliferation and Survival.

Normal myelopoiesis is regulated by the delicate interplay between hematopoietic stem cells (HSCs) and the bone marrow microenvironment. Through the action of hematopoietic growth factors and their receptors, alterations of transcriptional regulation result in myeloid progenitor cell proliferation and survival. Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF) and Interleukin-3 (IL-3) are cytokines that regulate myelopoiesis. GMCSF and IL-3 induce signaling pathways that activate serine threonine kinases, including MEK, ERK, and pp90RSK, leading to expression of immediate early genes such as early growth response gene-1 (egr-1). Analysis of promoter regions of egr-1 demonstrated that the cAMP response element, CRE, was necessary and sufficient for egr-1 expression in myeloid cells. We previously demonstrated that the transcription factor, CREB (cAMP Response Element Binding Protein), recognizes the CRE in the egr-1 promoter and is phosphorylated at serine 133 in response to GM-CSF and IL-3 through a MEK-dependent, protein kinase A-independent pathways. CREB is a leucine zipper transcription factor that regulates proliferation, differentiation, and survival of neuronal cells and lymphocytes. The majority of patients with acute lymphoid and myeloid leukemias express higher levels of CREB in their bone marrow. Therefore, we hypothesized that CREB plays a critical role in the regulation of normal hematopoiesis. To this end, we studied the biological and molecular effects of CREB in myeloid cells. CREB overexpression results in increased proliferation and survival through induction of cyclin A1 and other target genes. Transgenic mice that overexpress CREB in Gr-1/Mac-1+ cells develop myeloproliferative disease after one year. Knockdown of CREB using shRNAs leads to decreased proliferation and differentiation of myeloid progenitor cells and increased apoptosis of HSCs. Cell cycle analysis demonstrate that CREB downregulation in HSCs leads to greater numbers of cells in G1 and fewer cells in S phase. Both cyclin D1 and A1 levels are decreased in CREB shRNA-transduced HSCs. However, CREB knockdown of bone marrow progenitors did not affect long-term engraftment in transplantation assays. Recent findings on downstream pathways of CREB and potential target genes will be presented.