CXCR4 coordinates adhesion, migration, and development of human NK cells
Shira E. Eisman, Francesca E. Grossberg, Batya S. Koenigsberg, David H. McDermott, Frédérique van den Haak, Luis A. Pedroza, Everardo Hegewisch-Solloa, Philip M. Murphy, Emily M. Mace
Shira E. Eisman, Francesca E. Grossberg, Batya S. Koenigsberg, David H. McDermott, Frédérique van den Haak, Luis A. Pedroza, Everardo Hegewisch-Solloa, Philip M. Murphy, Emily M. Mace
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Research Article Cell biology Immunology

CXCR4 coordinates adhesion, migration, and development of human NK cells

Abstract

Natural killer (NK) cells undergo stepwise differentiation from multipotent progenitors within secondary lymphoid tissues. Despite the central importance of the tissue microenvironment in their development, little is known about cell-cell interactions that regulate human NK cell trafficking and maturation. Here, we identify the chemokine receptor CXCR4 and its ligand CXCL12 as regulators of stromal–NK cell interactions required for NK cell maturation. We demonstrate that CXCR4 is expressed throughout human NK cell development in peripheral blood and tonsil, and CXCL12 is enriched in stromal niches containing developing NK cells. Pharmacologic blockade or genetic disruption of CXCR4 resulted in diminished adhesion to integrin ligands, and high-resolution imaging demonstrated crosstalk between CXCR4 and integrins, providing a mechanistic basis for chemokine-dependent modulation of adhesion. Further, CXCR4 blockade resulted in altered contact-dependent motility on stromal cells and integrin ligands, with decreased stable stromal engagement and increased cell speed. Consistent with a requirement for these interactions, treatment with the CXCR4 antagonist plerixafor (AMD3100) impaired NK cell generation from CD34+ precursors. Analysis of NK cells from patients with WHIM syndrome with CXCR4 gain-of-function mutations treated with plerixafor revealed similar defects in migration and adhesion, supporting the in vivo relevance of CXCR4-dependent regulation of NK cell adhesion and motility.

Authors

Shira E. Eisman, Francesca E. Grossberg, Batya S. Koenigsberg, David H. McDermott, Frédérique van den Haak, Luis A. Pedroza, Everardo Hegewisch-Solloa, Philip M. Murphy, Emily M. Mace

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Figure 4

CXCR4 signaling promotes adhesion and integrin activation.

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CXCR4 signaling promotes adhesion and integrin activation. (A) Cell adhe...
(A) Cell adhesion assays of primary CD56+ NK cells on ICAM-1 following vehicle or 10 μM AMD3100 treatment. n = 4 donors; mean of each vehicle-treated donor is normalized to 1. (B) Adhesion of NK92 WT or CXCR4-KO cells to ICAM-1–coated plates. n = 3 independent experiments; large symbols represent the mean of each technical replicate. Statistical analysis by 2-tailed 1-sample t test compared with a theoretical mean of 1. (C) Surface CD18 expression in primary NK cells after treatment with vehicle (water) or AMD3100 displayed as representative histograms (left) and normalized MFI (right). Gray, fluorescence minus one (FMO) control. n = 3 independent experiments (mean ± SD). No statistical significance by 2-tailed Wilcoxon’s matched-pairs signed-rank test. (D) Representative confocal imaging of actin in NK92 cells treated with 10 μM of AMD3100 or vehicle and incubated on ICAM-1, then fixed and immunostained with phalloidin. Scale bars: 5 μm. Data are representative of 3 independent experiments/donors. (E) Primary CD56+ NK cells were treated with 10 μM of AMD3100 or vehicle and incubated on ICAM-1 (LFA-1 ligand) before fixation and immunostaining with anti–activated LFA-1 (clone m24), phalloidin, and DAPI. Scale bars: 5 μm; maximum projection shown. (F) Quantification of m24 MFI and percentage area from one representative donor. Each dot represents a single cell from one donor. Horizontal bars indicate mean ± SD. Statistical comparison by 2-tailed Mann-Whitney test across single-cell measurements within this donor. Data are representative of 3 independent experiments/donors (see also Supplemental Figure 5).