Cloning and characterization of a novel murine beta chemokine receptor, D6 - Comparison to three other related macrophage inflammatory protein-1 alpha receptors, CCR-1, CCR-3, and CCR-5

The beta-chemokine macrophage inflammatory protein-1 alpha (MIP-1 alpha) is chemotactic for many hemopoietic cell types and can inhibit hemopoietic stem cell (HSC) proliferation, effects mediated through G-protein coupled heptahelical receptors. We have isolated cDNAs for seven chemokine receptors, CCR-1 to -5, MIP-1 alpha RL1, and a novel cDNA, D6. Chinese hamster ovary cells expressing CCR-1, -3, -5, and D6 bound I-125-murine MIP-1 alpha: the order of affinity was D6 > CCR-5 > CCR-1 > CCR-3. Each bound a distinct subset of other beta-chemokines: the order of competition for I-125-murine MIP-1 alpha on D6 was murine MIP-1 alpha > human and murine MIP-1 beta > human RANTES similar to JE > human MCP-3 > human MCP-1. Human MIP-1 alpha and the alpha-chemokines did not compete. Like other chemokine receptors, He induced transient increases in [Ca2+] in HEK 293 cells upon ligand binding. D6 mRNA was abundant in lung and detectable in many other tissues, Bone marrow cell fractionation demonstrated T-cell and macrophage/monocyte expression of D6, and CCR-1, -3, and -5. Moreover, we could detect expression of CCR-3, CCR-5, and to a greater extent D6 in a cell population enriched for HSCs. Thus, we have characterized four murine beta chemokine receptors that are likely involved in mediating the pro-inflammatory functions of MIP-1 alpha and other chemokines, and we present D6, CCR-3, and CCR-5 as candidate receptors in MIP-1 alpha-induced HSC inhibition.