Granulocyte colony-stimulating factor rapidly activates a distinct STAT-like protein in normal myeloid cells.
Binding of granulocyte colony-stimulating factor (G-CSF) to normal myeloid cells activates the protein tyrosine kinases Lyn and Syk and results in the immediate early upregulation of G-CSF receptor (R) mRNA. In our studies of the signaling pathways activated by G-CSF that are coupled to proliferation and differentiation of myeloid cells, we examined whether G-CSF activated a latent transcription factor belonging to the STAT protein family. Electrophoretic mobility shift assays (EMSAs) of nuclear extracts from G-CSF-stimulated human myeloid cells showed the rapid activation of a DNA-binding protein that bound to the high-affinity serum-inducible element (hSIE) and migrated with mobility similar to serum inducible factor (SIF)-A (Stat3 homodimer). The G-CSF-stimulated SIF-A complex (G-SIF-A) did not bind to duplex oligonucleotides used to purify and characterize other Stat proteins (Stat1-6). In addition, antibodies raised against Stat1-6 failed to supershift the G-SIF-A complex or interfere with its formation. Based on its binding to the hSIE and lack of antigenic cross-reactivity with other known STAT proteins that bind to this element, it is likely that G-SIF-A is composed of a distinct member of the STAT protein family. EMSAs of whole-cell extracts prepared from cell lines containing full-length and truncated mutants of the G-CSFR showed that activation of G-SIF-A did not correlate with proliferation; rather, optimal activation requires the distal half of the cytosolic domain of the G-CSFR that is essential for differentiation. Activation of G-SIF-A, therefore, may be an early G-CSFR-coupled event that is critical for myeloid maturation.
Department of Medicine, University of Pittsburgh School of Medicine, PA, USA.
D J
DJ
Tweardy
T M
TM
Wright
S F
SF
Ziegler
H
H
Baumann
A
A
Chakraborty
S M
SM
White
K F
KF
Dyer
K A
KA
Rubin
D001402:B-Lymphocytes / Q000187:drug effects*; D001483:Base Sequence; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D004792:Enzyme Precursors / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects*; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology*; D006412:Hematopoietic Stem Cells / Q000187:drug effects*; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009504:Neutrophils / Q000187:drug effects*; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction / Q000502:physiology*; D000072377:Syk Kinase; D015534:Trans-Activators / Q000302:isolation & purification / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects; D019061:src-Family Kinases / Q000378:metabolism
D004268:DNA-Binding Proteins; D004792:Enzyme Precursors; D047908:Intracellular Signaling Peptides and Proteins; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D015534:Trans-Activators; D016179:Granulocyte Colony-Stimulating Factor; D007371:Interferon-gamma; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases