Research Project
10113925
Project Summary Cell fate specification in the hematopoietic system depends on the expression of lineage specific transcription factors. For instance, transcription factor C/EBP? promotes neutrophil development whereas high expression of PU.1 favors monocyte over neutrophil development. However, the roles of cytokines in cell fate determination are less clear and little is known about the intracellular signaling mechanisms by which cytokines instruct cell fate. G-CSF and M-CSF are two lineage-specific cytokines that play a dominant role in granulopoiesis and monopoiesis in response to G-CSF, respectively. Our preliminary studies suggest that prolonged Erk1/2 signaling led to monocyte development in response to G-CSF, which was associated with augmented activation of the ERK1/2 downstream targets c-Fos and Egr1. In contrast, suppression of ERK1/2 signaling resulted in predominantly neutrophil development in response to M-CSF. The expression of c-Fos and Egr1 was activated by PU.1, but repressed by C/EBP?. Interestingly, PU.1-mediated activation of c-Fos and Egr1 was markedly suppressed upon inhibition of ERK1/2 pathway, which was associated with a shift to neutrophil development in response to PU.1 activation. We hypothesize that lineage specific cytokines and transcription factors control neutrophil versus monocyte lineage choice in part through their differential effects on the ERK1/2-c-Fos/Egr1 pathway. Three specific aims are proposed to test the hypotheses. In aim 1, we will determine the role of ERK1/2 pathway in neutrophil versus monocyte lineage choice in response to G-CSF and M-CSF. We will address whether selective activation of ERK1/2 or overexpression of c-Fos and/or Egr1 is sufficient to cause monocyte development in response to G-CSF in primary common myeloid progenitors (CMPs) and granulocyte-monocyte progenitors (GMPs). Conversely, we will evaluate the effects of ERK1/2 inhibition and c-Fos/Egr1 knockdown on M-CSF-induced monocyte development in CMPs and GMPs. In aim 2, we will evaluate the effect of ERK1/2 inhibition on neutropenia resulting from loss of G-CSFR. We will investigate whether transgenic expression of a dominant negative (DN) MEK1 mutant, MEK1 K97M or oral administration of a FDA-approved MEK1/2 inhibitor trametinib alleviates neutropenia in CSF3R-/- mice. In aim 3, we will determine the role of ERK1/2-c-Fos/Egr1 pathway in PU.1-dependent monocyte development. We will determine whether PU.1 binds to the promoters of c-Fos and Egr1 to activate their transcription. We will also address whether inhibition of ERK1/2 signaling has an effect on PU.1-mediated monocyte development in CMPs and GMPs. The proposed research may reveal the key mechanisms by which lineage specific cytokines and transcription factors act in collaboration to regulate neutrophil versus monocyte lineage choice, and may lead to the development of novel therapeutic strategies for the treatment of neutropenia patients with CSF3R mutations.
