Increased phosphorylation of the colony stimulating factor-1 receptor following transmembrane signaling.
The receptor for colony-stimulating factor-1 (CSF-1) is a 165-kDa tyrosine kinase phosphorylated on serine residues in quiescent macrophages. Addition of CSF-1 to macrophages induces the following sequence of events: (1) formation of a 380-kDa noncovalently associated receptor dimer that is tyrosine phosphorylated; (2) protein tyrosine phosphorylation; (3) disulfide bond crosslinking of the noncovalently associated receptor monomers; and (4) conversion of the receptor homodimer to a heterodimeric 450-kDa species in which the apparent molecular mass of one of the subunits is increased from 165 to approximately 240 kDa (Li and Stanley, 1991). To study the kinetics of early phosphorylation/dephosphorylation events associated with these changes in the CSF-1R, we stimulated a CSF-1-dependent macrophage cell line with CSF-1 at 4 degrees C to slow reaction rates and prevent internalization. CSF-1 induced a rapid (2-4 min) initial increase in tyrosine phosphorylation of the 165-kDa subunits of the noncovalent 380-kDa homodimer. At 60-120 min after ligand binding, the conversion of some of the 165-kDa species to a approximately 240-kDa species on reducing SDS-PAGE was indicative of the transition of 380-kDa homodimer to 450-kDa heterodimer. Both the approximately 200 and 165-kDa subunits of the heterodimer were more highly phosphorylated in tyrosine than the subunits of the homodimer 2-4 min after ligand binding. The 165-kDa subunit of the heterodimer also exhibited increased phosphorylation in serine. However, in vitro dephosphorylation experiments indicated that a modification other than phosphorylation is involved in the conversion of the 165-kDa to the approximately 240-kDa CSF-1R species. The possibility that the increased tyrosine and serine phosphorylation of the subunits of the 450-kDa heterodimer may be involved in regulation of receptor kinase activity and/or internalization is discussed.
Baccarini
M
M
Department of Developmental Biology and Cancer, Albert Einstein College of Medicine, Bronx, NY 10461.
Li
W
W
Dello Sbarba
P
P
Stanley
E R
ER
D000469:Alkaline Phosphatase; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000502:physiology; D007700:Kinetics; D008970:Molecular Weight; D010449:Peptide Mapping; D010748:Phosphopeptides / Q000032:analysis; D010761:Phosphorus Radioisotopes; D010766:Phosphorylation; D011233:Precipitin Tests; D016186:Receptor, Macrophage Colony-Stimulating Factor / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*
D000596:Amino Acids; D010748:Phosphopeptides; D010761:Phosphorus Radioisotopes; D016186:Receptor, Macrophage Colony-Stimulating Factor; D000469:Alkaline Phosphatase