CXCR7 Controls Competition for Recruitment of β-Arrestin 2 in Cells Expressing Both CXCR4 and CXCR7

Abstract

Chemokine CXCL12 promotes growth and metastasis of more than 20 different human cancers, as well as pathogenesis of other common diseases. CXCL12 binds two different receptors, CXCR4 and CXCR7, both of which recruit and signal through the cytosolic adapter protein β-arrestin 2. Differences in CXCL12-dependent recruitment of β-arrestin 2 in cells expressing one or both receptors remain poorly defined. To quantitatively investigate parameters controlling association of β-arrestin 2 with CXCR4 or CXCR7 in cells co-expressing both receptors, we used a systems biology approach combining real-time, multi-spectral luciferase complementation imaging with computational modeling. Cells expressing only CXCR4 maintain low basal association with β-arrestin 2, and CXCL12 induces a rapid, transient increase in this interaction. In contrast, cells expressing only CXCR7 have higher basal association with β-arrestin 2 and exhibit more gradual, prolonged recruitment of β-arrestin 2 in response to CXCL12. We developed and fit a data-driven computational model for association of either CXCR4 or CXCR7 with β-arrestin 2 in cells expressing only one type of receptor. We then experimentally validated model predictions that co-expression of CXCR4 and CXCR7 on the same cell substantially decreases both the magnitude and duration of CXCL12-regulated recruitment of β-arrestin 2 to CXCR4. Co-expression of both receptors on the same cell only minimally alters recruitment of β-arrestin 2 to CXCR7. In silico experiments also identified β-arrestin 2 as a limiting factor in cells expressing both receptors, establishing that CXCR7 wins the “competition” with CXCR4 for CXCL12 and recruitment of β-arrestin 2. These results reveal how competition for β-arrestin 2 controls integrated responses to CXCL12 in cells expressing both CXCR4 and CXCR7. These results advance understanding of normal and pathologic functions of CXCL12, which is critical for developing effective strategies to target these pathways therapeutically.

Macrophage Migration Inhibitory Factor-CXCR4 Receptor Interactions

“…The P1V/M2S mutant did not activate CXCR4, suggesting that the MIF catalytic cavity is important for binding or inducing CXCR4 signaling. A dose-response effect using extracellular MIF is observed, but an EC 50 value cannot be obtained because a plateau cannot be reached at the higher concentrations. In comparison, exogenous CXCL12 has about a 100-fold increase of the EC 50 value in S. cerevisiae relative to mammalian cells (data not shown).…”

“…D, dose-response effect of exogenous MIF added to CXCR4-expressing S. cerevisiae. The EC 50 values cannot be measured because a concentration that reaches the maximum signaling cannot be obtained. E, functional competition between MIF and CXCL12 in activating CXCR4.…”

“…MIF did not activate ␤-arrestin 2, whereas CXCL12 showed significant ␤-arrestin 2 activation (Fig. 7A) (50). The lack of any interaction is surprising given results from Jurkat T-cells that show 40% CXCR4 internalization by MIF (whereas CXCL12 results in 80% internalization), presumably induced by ␤-arrestin 2 (37).…”

“…Data were graphed as mean values Ϯ S.E. for fold change in bioluminescence relative to vehicle control at each time point to account for dynamics of the luciferin-luciferase reaction (n ϭ 4 per condition) (50). We present data representative of two independent experiments.…”

Chemokine signaling in development and disease

“…As CXCR4 and CXCR7 both interact with β-arrestins, the relative levels of these two receptors in a cell can influence whether downstream signaling is preferentially mediated through β-arrestinlinked pathways or through G protein pathways (Coggins et al, 2014;Decaillot et al, 2011;Sierro et al, 2007). Moreover, CXCL12 can bind to CXCR4 as a monomer or as a dimer in vitro.…”