Multiple signaling pathways induced by granulocyte colony-stimulating factor involving activation of JAKs, STAT5, and/or STAT3 are required for regulation of three distinct classes of immediate early genes.
Granulocyte colony-stimulating factor (G-CSF) is the major regulator of proliferation and differentiation of neutrophilic granulocyte precursor cells. G-CSF activates multiple signaling molecules, including the JAK1 and JAK2 kinases and the STAT transcription factors. To investigate G-CSF signaling events regulated by the JAK-STAT pathway, we have generated UT7-epo cells stably expressing either wild-type (wt) G-CSF receptor or a series of C-terminal deletion mutants. Gel mobility shift and immunoprecipitation/Western analysis showed that STAT5 is rapidly activated by G-CSF in cells expressing the wt G-CSF receptor, in addition to the previously reported STAT3 and STAT1. Mutants lacking any tyrosine residues in the cytoplasmic domain maintain their ability to activate STAT5 and STAT1 but cannot activate STAT3, implying that STAT5 and STAT1 activation does not require receptor tyrosine phosphorylation. We also observed significant changes in the ratio of STAT1:STAT3:STAT5 activated by various G-CSF receptor C-terminal deletion mutants. These mutant receptors were further used to investigate the role of JAKs and STATs in G-CSF-mediated responses in these cells. We found that JAK activation correlates with G-CSF-induced cell proliferation, whereas STAT activation is not required. We have also identified three classes of G-CSF immediate early genes, whose activation correlates with the activation of distinct JAK-STAT pathways. Our data show that, whereas c-fos is regulated through a pathway independent of STAT activation, oncostatin M, IRF-1, and egr-1 are regulated by an STAT5-dependent pathway and fibrinogen is regulated by an STAT3-dependent pathway. In conclusion, our results suggest that G-CSF regulates its complex biologic activities by selectively activating distinct early response genes through different JAK-STAT signaling molecules.
Tian
S S
SS
Ligand Pharmaceuticals, San Diego, CA 92121, USA.
Tapley
P
P
Sincich
C
C
Stein
R B
RB
Rosen
J
J
Lamb
P
P
D000818:Animals; D002455:Cell Division / Q000235:genetics; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000502:physiology*; D004789:Enzyme Activation / Q000502:physiology; D005786:Gene Expression Regulation; D017781:Genes, Immediate-Early / Q000235:genetics*; D016179:Granulocyte Colony-Stimulating Factor / Q000235:genetics / Q000494:pharmacology*; D006801:Humans; D053616:Janus Kinase 3; D008894:Milk Proteins; D009154:Mutation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D050796:STAT3 Transcription Factor; D050799:STAT5 Transcription Factor; D015398:Signal Transduction / Q000187:drug effects* / Q000235:genetics; D015534:Trans-Activators / Q000502:physiology*
D004268:DNA-Binding Proteins; D008894:Milk Proteins; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; D050799:STAT5 Transcription Factor; D015534:Trans-Activators; D016179:Granulocyte Colony-Stimulating Factor; D011505:Protein-Tyrosine Kinases; C507933:JAK3 protein, human; D053616:Janus Kinase 3