Abstract
Chemokine-driven migration is accompanied by polarization of the cell body and of the intracellular signaling machinery. The extent to which chemokine receptors polarize during chemotaxis is currently unclear. To analyze the distribution of the chemokine receptor CXCR4 during SDF-1 (CXCL12)-induced chemotaxis, we retrovirally expressed a CXCR4-GFP fusion protein in the CXCR4-deficient human hematopoietic progenitor cell line KG1a. This KG1a CXCR4-GFP cell line showed full restoration of SDF-1 responsiveness in assays detecting activation of ERK1/2 phosphorylation, actin polymerization, adhesion to endothelium under conditions of physiological flow, and (transendothelial) chemotaxis. When adhered to cytokine-activated endothelium in the absence of SDF-1, CXCR4 did not localize to the leading edge of the cell but was uniformly distributed over the plasma membrane. In contrast, when SDF-1 was immobilized on cytokine-activated endothelium, the CXCR4-GFP receptors that were present on the cell surface markedly redistributed to the leading edge of migrating cells. In addition, CXCR4-GFP co-localized with lipid rafts in the leading edge of SDF-1-stimulated cells, at the sites of contact with the endothelial surface. Inhibition of lipid raft formation prevents SDF-1-dependent migration, internalization of CXCR4, and polarization to the leading edge of CXCR4, indicating that CXCR4 surface expression and signaling requires lipid rafts. These data show that SDF-1, immobilized on activated human endothelium, induces polarization of CXCR4 to the leading edge of migrating cells, revealing co-operativity between chemokine and substrate in the control of cell migration.
Highlights
IntroductionIn the process of homing, transplanted CD34ϩ cells migrate from the peripheral blood across the endothelium back to their niche in the bone marrow
It has been shown that SDF-1 stimulates integrin-mediated arrest of CD34ϩ cells and of T lymphocytes on vascular endothelium under flow, and that SDF-1 is constitutively expressed on human bone marrow endothelium [2, 7]
Directional migration of leukocytes is accompanied by polarization of the cell body and of the actin cytoskeleton, redistribution of chemokine receptors has been the subject of contradictory reports
Summary
In the process of homing, transplanted CD34ϩ cells migrate from the peripheral blood across the endothelium back to their niche in the bone marrow. Recent evidence indicates that differential distribution of G␥-subunits leads to amplification of the chemokine receptor signaling [11]. It has been reported that SDF-1 can bind to fibronectin and subsequently induces a polarized distribution of CXCR4 on adherent T lymphocytes [15]. These studies were all performed with artificial substrates, and to our knowledge it is unclear whether leukocytes that adhere to activated endothelial cells redistribute chemokine receptors, such as. There is no information on the distribution and localization of CXCR4 in human hematopoietic progenitor cells
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