GRANULOCYTE-COLONY-STIMULATING FACTOR-RECEPTOR SIGNALING INVOLVES THEFORMATION OF A 3-COMPONENT COMPLEX WITH LYN AND SYK PROTEIN-TYROSINE KINASES

Granulocyte colony-stimulating factor (G-CSF) is a glycoprotein that c ritically regulates the viability, proliferation, and differentiation of granulocytic precursors and the function of neutrophils by signalin g through its receptor. Cloning of the human G-CSF receptor (G-CSFR) c DNA has demonstrated sequence homology with other members of the hemat opoietic/cytokine receptor superfamily. G-CSF stimulates the appearanc e of phosphotyrosine proteins in several types of human and murine mye loid cells. Since the receptor does not possess intrinsic tyrosine kin ase activity, we hypothesized that G-CSFR interacts with and activates cytosolic protein-tyrosine kinases (PTKs). In vitro protein kinase as say of human G-CSFR immunoprecipitates demonstrated at least two tyros ine phosphoproteins, pp55 and pp70. We observed that G-CSF activated p 53/p56lyn, a Src-related PTK, and p72syk, a non-Src-related PTK. Lyn a nd Syk were recovered in anti-G-CSFR immunoprecipitates; Lyn was detec ted in the absence of ligand. In addition, upon G-CSF stimulation, Lyn coimmunoprecipitated with Syk. Analysis of the G-CSFR amino acid sequ ence revealed a potential receptor activation motif for Syk. On the ba sis of immunoprecipitation and sequence analysis data, we propose that the human G-CSFR forms a three-component signaling complex with Lyn a nd Syk. Their sequential recruitment into the G-CSFR signaling complex demonstrates the coordinated involvement of two PTKs with a member of the hematopoietic/cytokine receptor superfamily.