The Duration of Erk1/2 Signaling Determines Neutrophil Versus Monocyte Cell Fates in Response to G-CSF and M-CSF

Neutrophils and monocytes/macrophages are derived from hematopoietic stem cells that, through progressive commitment, give rise to granulocyte-monocyte progenitors that in turn develop into either neutrophils or monocytes/macrophages. Although it is well known that cell fate specification in the hematopoietic system depends on the expression of lineage specific transcription factors, the roles of cytokines in lineage commitment are less clear and two models have been proposed. According to the stochastic model, cell fate choice is stochastic and cytokines simply provide signals for the survival and proliferation of committed cells. The instructive model, on the other hand, proposes that cytokines stimulate intracellular signaling pathways that dictate cell fate decisions. G-CSF and M-CSF are two lineage-specific cytokines that play a dominant role in granulopoiesis and monopoiesis, respectively. Recent studies lend strong support to the roles of G-CSF and M-CSF in instructing lineage commitment. However, the signaling pathways that determine neutrophil versus monocyte cell fate following stimulation with G-CSF and M-CSF are unknown. Here we show that tyrosine (Y) 729 of the G-CSFR is involved in transducing signals that specify neutrophil cell fate. Substitution of Y729 with phenylalanine (F) results in monocytic differentiation in response to G-CSF in murine myeloid 32D and multipotent FDCP-mix A4 cells. G-CSF stimulated activation of Erk1/2 was prolonged in cells expressing G-CSFR Y729F mutant. Significantly, treatment of cells with Mek1/2 inhibitors U0126 or PD0325901 rescued neutrophilic differentiation. M-CSF has been shown to induce prolonged activation of Erk1/2, which is required for monocytic differentiation. Interestingly, the Mek1/2 inhibitors also promoted neutrophil cell fate at the expense of monocytic development in lineage marker negative (Lin^-) primary bone marrow cells cultured in M-CSF. We further demonstrate that prolonged activation of Erk1/2 was associated with augmented activation of c-Fos and Egr1, both of which have previously been shown to promote monocytic development. Consistent with this, knockdown of c-Fos or Egr1 redirected 32D cells expressing G-CSFR Y729F mutant to develop into neutrophils in response to G-CSF. We propose that M-CSF stimulates more sustained activation of Erk1/2 than G-CSF does and that the duration of Erk1/2 signaling regulates neutrophil versus monocyte cell fate choices, likely through altering the activation statuses of c-Fos and Egr1.