The Src-like Adaptor Protein 2 Regulates Colony-stimulating Factor-1 Receptor Signaling and Down-regulation

Abstract

The colony-stimulating factor 1 receptor 4 is a member of the type III receptor tyrosine kinase (RTK) family, which includes c-Kit, platelet-derived growth factor receptor ␣ and ␤, and Flt3 (1). All of the functions of colony-stimulating factor-1 (CSF-1) are mediated by CSF-1R, which is the primary regulator of the common myeloid lineage consisting of osteoclasts and mononuclear phagocytes (monocytes and macrophages) in vivo (2-5).Ligand binding induces CSF-1R dimerization and autophosphorylation on tyrosine residues that regulate both kinase activity and mediate binding to downstream SH2 and PTB domain-containing proteins. Recruitment of such proteins leads to activation of signaling cascades involved in determining the biological outcomes of CSF-1R activation as well as receptor down-regulation (6 -8).A number of adaptor proteins, including Gads/Mona, FMIP, DOK-2, and c-Cbl, have been implicated as negative regulators of CSF-1R signaling. c-Cbl possesses a variant SH2 domain (TKB domain) that mediates binding to activated tyrosine kinases and a RING finger domain demonstrated to confer E3 ubiquitin ligase activity, which promotes the ubiquitination of activated tyrosine kinases (9, 10). In c-Cbl-deficient macrophages, both CSF-1R ubiquitination and endocytosis of the ubiquitinated receptor are impaired (11). Subsequent to CSF-1R ubiquitination, activated CSF-1R is degraded intralysosomally (12). The degradation of ligand-stimulated CSF-1R normally occurs rapidly, and the half-life of the CSF-1R decreases from 3 h to 5 min upon ligand binding (13,14). Mutation of the tyrosine residue within the c-Cbl binding site (Tyr-969 in human CSF-1R) has been frequently observed in myelodysplasia and acute myeloblastic leukemia (15). Similarly, the transforming viral homologue of CSF-1R, v-Fms, lacks the c-Cbl binding site. Importantly, re-addition of the c-Cbl binding site to v-Fms decreases its transforming activity (16).Src-like adaptor protein family proteins, SLAP and SLAP-2, have an amino-terminal myristoylation signal required for association with cell membranes, closely juxtaposed SH3 and SH2 domains, and a unique carboxyl-terminal region that mediates association with c-Cbl. Both SLAP and SLAP-2 have been demonstrated to function as negative regulators of T-cell antigen receptor (TCR) signaling via a mechanism requiring the c-Cbl binding region (17)(18)(19). In addition to c-Cbl, SLAP-2 associates with the Zap-70 and Syk cytoplasmic tyrosine kinases and promotes their degradation when co-expressed. Recent work has shown that SLAP functions as a negative regulator of the TCR by promoting c-Cbl-dependent ubiquitination of TCR chain and down-regulation of the CD3 complex (17,20,21). In addition, both SLAP and SLAP-2 have been implicated as negative regulators of B-cell receptor signaling (19,22), and avian SLAP has been demonstrated to interfere with erythropoietin signaling in erythroblasts (23).Although SLAP and SLAP-2 have been implicated in the regulation of antigen receptor signaling, SLAP was originally dis-* ...

The Src-Like Adaptor Protein Regulates GM-CSFR Signaling and Monocytic Dendritic Cell Maturation

“…The ability of SLAP to be induced under particular circumstances suggests that it may regulate multiple receptors and pathways. Previous studies from our group and other investigators showed that SLAP2 binds to and regulates M-CSFR and CSF-1R; expression of dominant negative SLAP2 results in a potentiation of CSF-1-mediated differentiation (14,16). Similar to the situation in T cells, these effects also seem to be mediated through c-Cbl (14).…”

“…Although many SLAP-and SLAP2-binding partners have been mapped to the SH2 domain (9)(10)(11)(12)(13)(14), we found that SLAP interacted with the a subunit of GM-CSFR, which, in the mouse, does not contain any potential tyrosine-phosphorylation sites within the cytoplasmic tail. However, GM-CSFRa contains a PXXP motif that may mediate SH3 domain interactions (34,35).…”

“…Previous studies from our group and other investigators showed that SLAP2 binds to and regulates M-CSFR and CSF-1R; expression of dominant negative SLAP2 results in a potentiation of CSF-1-mediated differentiation (14,16). Similar to the situation in T cells, these effects also seem to be mediated through c-Cbl (14). In this study, we examined the role of SLAP and SLAP2 in cytokine receptor-signaling pathways by studying GM-CSF responses in primary BM-DC.…”

“…SLAP and SLAP2 are expressed in many hematopoietic cell types and have a negative regulatory role in the regulation of TCR, BCR, and CSF-1R (8)(9)(10)(11)(12)(13)(14)(15)(16). Our previous studies showed a role for SLAP2 in CSF-1R signaling, leading us to examine whether SLAP and SLAP2 could be involved in the regulation of hematopoietic cytokine signaling and early myeloid differentiation.…”

“…TCR regulation by SLAP is important during development, because T cells from SLAP 2/2 mice display defects in the double-positive stage (CD4 + CD8 + ) of thymocyte development and not in peripheral T cells (9). Although SLAP and SLAP2 seem to play a role in T cell signaling, their expression is not limited to T cells; they are expressed in a variety of hematopoietic cell types, including B cells, mast cells, macrophages, and early progenitor cells (13)(14)(15)(16).…”