title: Beta-chemokine receptor CCR5 signals through SHP1, SHP2, and Syk.
abstract: The beta-chemokine receptor CCR5 has been shown to modulate cell migration, proliferation, and immune functions and to serve as a co-receptor for the human immunodeficiency virus. We and others have shown that CCR5 activates related adhesion focal tyrosine kinase (RAFTK)/Pyk2/CAK-beta. In this study, we further characterize the signaling molecules activated by CCR5 upon binding to its cognate ligand, macrophage inflammatory protein-1beta (MIP1beta). We observed enhanced tyrosine phosphorylation of the phosphatases SHP1 and SHP2 upon MIP1beta stimulation of CCR5 L1.2 transfectants and T-cells derived from peripheral blood mononuclear cells. Furthermore, we observed that SHP1 associated with RAFTK. However, using a dominant-negative phosphatase-binding mutant of RAFTK (RAFTK(m906)), we found that RAFTK does not mediate SHP1 or SHP2 phosphorylation. SHP1 and SHP2 also associated with the adaptor protein Grb2 and the Src-related kinase Syk. Pretreatment of CCR5 L1.2 transfectants or T-cells with the phosphatase inhibitor orthovanadate markedly abolished MIP1beta-induced chemotaxis. Syk was also activated upon MIP1beta stimulation of CCR5 L1.2 transfectants or T-cells and associated with RAFTK. Overexpression of a dominant-negative Src-binding mutant of RAFTK (RAFTK(m402)) significantly attenuated Syk activation, whereas overexpression of wild-type RAFTK enhanced Syk activity, indicating that RAFTK acts upstream of CCR5-mediated Syk activation. Taken together, these results suggest that MIP1beta stimulation mediated by CCR5 induces the formation of a signaling complex consisting of RAFTK, Syk, SHP1, and Grb2.
authors.affiliation: Division of Experimental Medicine, Hematology/Oncology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts 02115, USA. rganju@caregroup.harvard.edu
authors.forename: R K
authors.initials: RK
authors.lastname: Ganju
authors.forename: S A
authors.initials: SA
authors.lastname: Brubaker
authors.forename: R D
authors.initials: RD
authors.lastname: Chernock
authors.forename: S
authors.initials: S
authors.lastname: Avraham
authors.forename: J E
authors.initials: JE
authors.lastname: Groopman
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D054407:Chemokine CCL4; D002634:Chemotaxis, Leukocyte / Q000187:drug effects / Q000502:physiology; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004792:Enzyme Precursors / Q000378:metabolism*; D051418:Focal Adhesion Kinase 2; D051380:GRB2 Adaptor Protein; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D016393:Lymphoma, B-Cell; D019402:Macrophage Inflammatory Proteins / Q000494:pharmacology; D016296:Mutagenesis; D010766:Phosphorylation; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D019713:Receptors, CCR5 / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction; D000072377:Syk Kinase; D013601:T-Lymphocytes / Q000502:physiology; D014162:Transfection; D014407:Tumor Cells, Cultured; D014638:Vanadates / Q000494:pharmacology
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D054407:Chemokine CCL4; D004791:Enzyme Inhibitors; D004792:Enzyme Precursors; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D019402:Macrophage Inflammatory Proteins; D011506:Proteins; D019713:Receptors, CCR5; D011994:Recombinant Proteins; D014638:Vanadates; D011505:Protein-Tyrosine Kinases; D051418:Focal Adhesion Kinase 2; C000606896:SYK protein, human; D000072377:Syk Kinase; C516724:PTPN11 protein, human; C509249:PTPN6 protein, human; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases