The CXCR4 agonist ligand stromal derived factor-1 maintains high affinity for receptors in both G(alpha i)-coupled and uncoupled states

The alpha chemokine receptor CXCR4 and its only characterized chemokine lig and, stromal cell-derived factor-1 (SDF-1), are postulated to be important in the development of the B-cell arm of the immune system. In addition, CXC R4 is a critical coreceptor in support of viral entry by T-cell line tropic strains (X4) of the Human Immunodeficiency Virus Type 1 (HIV-1), viral var iants which predominate in some infected individuals in end stage disease. SDF1 can block X4-tropic HIV-1 infection of CD4 + target cells in vitro, an d allelic variants of the human gene encoding SDF-1 in vivo correlate with delayed disease progression. Therefore, CXCR4 may be an appropriate target for therapeutic intervention in acquired immunodeficiency syndrome (AIDS), and knowledge of the pharmacology of SDF-1 binding to its cognate receptor will be important in the interpretation of these experiments. We report her e a K-d derived using a competition binding assay of 4.5 nM for CXCR4 endog enously expressed on peripheral blood monocytes and T-cells. This affinity is similar to that which SDF-1 exhibits when binding to endogenous CXCR4 on an established immortal Jurkat T-cell line as well as recombinant CXCR4 tr ansfected into Chinese Hamster Ovary (CHO) cells. We also demonstrate that the determined affinity of SDF-1 for CXCR4 is reflective of its ability to induce a CXCR4-mediated signal transduction in these different cell types. Furthermore, using Bordetella pertussis toxin, we observe that high affinit y binding of SDF-1 to CXCR4 is independent of the G-protein coupled state o f the receptor, as uncoupling of G-protein did not lead to the appearance o f measurable low affinity SDF-1 binding sites. Moreover, binding affinity a nd receptor number were unaffected by uncoupling for both recombinant and e ndogenously expressed CXCR4. Thus, SDF-1 is novel among agonist ligands of G protein-coupled receptors in that it appears to have equal affinity for b oth the G protein-coupled and uncoupled states of CXCR4. (C) 2000 Elsevier Science B.V. All rights reserved.