The Low-Molecular-Weight Phosphotyrosine Protein Phosphatase, when Overexpressed, Reduces the Mitogenic Response to Macrophage Colony-Stimulating Factor and Tyrosine Phosphorylation of Its Receptor

1998 ◽  
Vol 253 (2) ◽  
pp. 300-304 ◽  
Author(s):  
Elisabetta Rovida ◽  
Stefania Rigacci ◽  
Alessandro Paccagnini ◽  
Persio Dello Sbarba ◽  
Andrea Berti
Keyword(s):  
Molecular Weight ◽  
Tyrosine Phosphorylation ◽  
Protein Phosphatase ◽  
Low Molecular Weight ◽  
Colony Stimulating Factor ◽  
Mitogenic Response ◽  
Macrophage Colony Stimulating Factor ◽  
Phosphotyrosine Protein Phosphatase ◽  
Macrophage Colony ◽  
Stimulating Factor

Granulocyte-macrophage colony-stimulating factor preferentially activates the 94-kD STAT5A and an 80-kD STAT5A isoform in human peripheral blood monocytes

Blood ◽  
1996 ◽  
Vol 88 (4) ◽  
pp. 1206-1214 ◽  
Author(s):  
RL Rosen ◽  
KD Winestock ◽  
G Chen ◽  
X Liu ◽  
L Hennighausen ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Human Peripheral Blood ◽  
Janus Kinase ◽  
Lower Molecular Weight ◽  
Colony Stimulating Factor ◽  
Enhancer Element ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Granulocyte-macrophage colony-stimulating factor (GM-CSF) induces immediate effects in monocytes by activation of the Janus kinase (JAK2) and STAT transcription factor (STAT5) pathway. Recent studies have identified homologues of STAT5, STAT5A, and STAT5B, as well as lower molecular weight variants of STAT5. To define the activation of the STAT5 homologues and lower molecular weight variant in human monocytes and monocytes differentiated into macrophages by culture in macrophage- CSF (M-CSF), we measured the GM-CSF induced tyrosine phosphorylation of STAT5A, STAT5B, and any lower molecular weight STAT5 isoforms. Freshly isolated monocytes expressed 94-kD STAT5A, 92-kD STAT5B, and an 80-kD STAT5A molecule. Whereas 94-kD STAT5A was clearly tyrosine phosphorylated and bound to the enhancer element, the gamma response region (GRR), of the Fc gamma RI gene, substantially less tyrosine phosphorylated STAT5B bound to the immobilized GRR element. Macrophages lost their ability to express the 80-kD STAT5A protein, but retained their ability to activate STAT5A. STAT5A-STAT5A homodimers and STAT5A- STAT5B heterodimers formed in response to GM-CSF. Therefore, activation of STAT5A predominates compared to STAT5B when assayed by direct immunoprecipitation and by evaluation of bound STATs to immobilized GRR. Selective activation of STAT5 homologues in addition to generation of lower molecular isoforms may provide specificity and control to genes expressed in response to cytokines such as GM-CSF.

Saturation Mutagenesis of the β Subunit of the Human Granulocyte-Macrophage Colony-Stimulating Factor Receptor Shows Clustering of Constitutive Mutations, Activation of ERK MAP Kinase and STAT Pathways, and Differential β Subunit Tyrosine Phosphorylation

Blood ◽  
1998 ◽  
Vol 92 (6) ◽  
pp. 1989-2002 ◽  
Author(s):  
Brendan J. Jenkins ◽  
Timothy J. Blake ◽  
Thomas J. Gonda
Keyword(s):  
Map Kinase ◽  
Tyrosine Phosphorylation ◽  
Transmembrane Domain ◽  
Point Mutations ◽  
Saturation Mutagenesis ◽  
Β Subunit ◽  
Colony Stimulating Factor ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract The high-affinity receptors for human granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and IL-5 are heterodimeric complexes consisting of cytokine-specific  subunits and a common signal-transducing β subunit (hβc). We have previously demonstrated the oncogenic potential of this group of receptors by identifying constitutively activating point mutations in the extracellular and transmembrane domains of hβc. We report here a comprehensive screen of the entire hβc molecule that has led to the identification of additional constitutive point mutations by virtue of their ability to confer factor independence on murine FDC-P1 cells. These mutations were clustered exclusively in a central region of hβc that encompasses the extracellular membrane-proximal domain, transmembrane domain, and membrane-proximal region of the cytoplasmic domain. Interestingly, most hβc mutants exhibited cell type-specific constitutive activity, with only two transmembrane domain mutants able to confer factor independence on both murine FDC-P1 and BAF-B03 cells. Examination of the biochemical properties of these mutants in FDC-P1 cells indicated that MAP kinase (ERK1/2), STAT, and JAK2 signaling molecules were constitutively activated. In contrast, only some of the mutant β subunits were constitutively tyrosine phosphorylated. Taken together, these results highlight key regions involved in hβc activation, dissociate hβc tyrosine phosphorylation from MAP kinase and STAT activation, and suggest the involvement of distinct mechanisms by which proliferative signals can be generated by hβc. © 1998 by The American Society of Hematology.

scholarly journals Macrophage colony-stimulating factor-induced tyrosine phosphorylation of c-fms proteins expressed in FDC-P1 and BALB/c 3T3 cells.

1990 ◽  
Vol 10 (6) ◽  
pp. 2528-2538 ◽  
Author(s):  
P Tapley ◽  
A Kazlauskas ◽  
J A Cooper ◽  
L R Rohrschneider
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Phosphorylation ◽  
Phosphorylation Sites ◽  
Colony Stimulating Factor ◽  
Major Site ◽  
Tryptic Peptides ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor ◽  
Autophosphorylation Site

The c-fms protein is a receptor for macrophage colony-stimulating factor (M-CSF) with intrinsic protein-tyrosine kinase activity. We investigated the tyrosine phosphorylation of murine c-fms proteins expressed from a retroviral vector in factor-dependent myeloid FDC-P1 cells and in BALB/c 3T3 fibroblasts transformed by the expression of the c-fms gene. FDC-P1 cells expressing c-fms were able to grow and differentiate in response to M-CSF. Their c-fms proteins were normally phosphorylated on serine and became phosphorylated on tyrosine residues contained in five tryptic peptides when the cells were exposed to M-CSF. A subset of these peptides was constitutively phosphorylated in BALB/c cells expressing c-fms, consistent with the production of M-CSF by these cells. All the peptides detected in vivo were also phosphorylated in vitro. These peptides were analyzed by susceptibility to proteases, comparison with synthetic peptides, and site-directed mutagenesis. The identities of four of the tryptic peptides were determined; they arise from three unique tyrosine phosphorylation sites. One major site of tyrosine phosphorylation at residue 697 accounted for two of the tryptic peptides. A second major site was identified at tyrosine residue 706. These two tyrosine phosphorylation sites are located within the tyrosine kinase insert region. Tyrosine 807, which has homology to the major autophosphorylation site of the p60v-src tyrosine kinase, is a minor autophosphorylation site. Possible functional roles for these phosphorylations of the c-fms protein include interactions with substrate proteins, catalytic activity, and ligand-induced degradation.

scholarly journals Hck expression correlates with granulocyte-macrophage colony- stimulating factor-induced proliferation in HL-60 cells

Blood ◽  
1994 ◽  
Vol 84 (1) ◽  
pp. 94-103
Author(s):  
D Linnekin ◽  
OM Howard ◽  
L Park ◽  
W Farrar ◽  
D Ferris ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Cell Line ◽  
Tyrosine Phosphorylation ◽  
Protein Tyrosine Phosphorylation ◽  
Colony Stimulating Factor ◽  
Protein Tyrosine ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

The human myeloid cell line HL-60 expresses approximately 300 high- affinity granulocyte-macrophage colony-stimulating factor receptors (GM- CSFRs), yet treatment of these cells with GM-CSF does not result in enhanced cellular proliferation or increases in protein tyrosine phosphorylation. In contrast, GM-CSF induces rapid increases in protein tyrosine phosphorylation and proliferative responses in HL-60 cells pretreated for 3 days in dimethyl sulfoxide (DMSO). Similarly, HL-60 cells pretreated with retinoic acid or 1,25 dihydroxyvitamin D3 were also capable of responding to GM-CSF. Interestingly, each of these treatments resulted in increased expression of the src-like tyrosine kinase hck. Stimulation with GM-CSF increased hck autophosphorylation in DMSO-treated HL-60 cells, suggesting that hck is a component of the GM-CSF signal transduction pathway. To determine if hck has a role in the DMSO-induced recoupling of the GM-CSFR, we overexpressed hck in HL- 60 cells. The resulting cell line (HL-60/hck) expresses hck mRNA and protein at levels comparable with DMSO-treated HL-60 cells. Stimulation of HL-60/hck cells with GM-CSF results in activation of hck, increases in protein tyrosine phosphorylation, and increased proliferation. These results show that cytokine receptors can exist in an uncoupled form and suggest that in HL-60 cells, appropriate levels of the src-like tyrosine kinase hck are critical for functional coupling of the GM-CSFR to biologic responses.

scholarly journals Granulocyte-macrophage colony-stimulating factor activates microtubule- associated protein 2 kinase in neutrophils via a tyrosine kinase- dependent pathway

Blood ◽  
1992 ◽  
Vol 79 (12) ◽  
pp. 3350-3354 ◽  
Author(s):  
MA Raines ◽  
DW Golde ◽  
M Daeipour ◽  
AE Nel
Keyword(s):  
Signal Transduction ◽  
Tyrosine Kinase ◽  
Tyrosine Phosphorylation ◽  
Kinase Activity ◽  
Colony Stimulating Factor ◽  
Kinase Activation ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Receptors of the hematopoietin superfamily, including the granulocyte- macrophage colony-stimulating factor (GM-CSF) receptor, lack a tyrosine kinase domain as well as other sequences indicative of a known signaling mechanism. In this report, we identify the serine/threonine kinase, microtubule-associated protein 2 (MAP2) kinase, as an intermediate in the GM-CSF signal transduction pathway. Treatment of peripheral blood neutrophils or terminally differentiated HL-60 cells with GM-CSF induced a rapid and dose-dependent increase in MAP2 kinase activity. Maximal activity occurred within 5 minutes and the kinetics of the response varied depending on the target cell (prolonged in neutrophils and transient in neutrophilic HL-60 cells). MAP2 kinase activity in these cells correlates with the induction of a 42-Kd tyrosine phosphoprotein. Furthermore, tyrosine phosphorylation is necessary for MAP2 kinase activation since its activity is inhibited by treatment with the tyrosine kinase inhibitor, erbstatin analog. These data suggest that tyrosine phosphorylation is important in GM-CSF- mediated signal transduction and that MAP2 kinase activation may be a central biochemical event involved in its signaling.

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