Functional Interaction of CD154 Protein with α5β1 Integrin Is Totally Independent from Its Binding to αIIbβ3 Integrin and CD40 Molecules

Youssef El Fakhry,Haydar Alturaihi,Daniel Yacoub,Lihui Liu,Wenyan Guo,Claire Leveillé,Daniel Jung,Lara Bou Khzam,Yahye Merhi,John A Wilkins,Hongmin Li,Walid Mourad

doi:10.1074/jbc.m111.333989

Abstract

In addition to its classical CD40 receptor, CD154 also binds to αIIbβ3, α5β1, and αMβ2 integrins. Binding of CD154 to these receptors seems to play a key role in the pathogenic processes of chronic inflammation. This investigation was aimed at analyzing the functional interaction of CD154 with CD40, αIIbβ3, and α5β1 receptors. We found that the binding affinity of CD154 for αIIbβ3 is ∼4-fold higher than for α5β1. We also describe the generation of sCD154 mutants that lost their ability to bind CD40 or αIIbβ3 and show that CD154 residues involved in its binding to CD40 or αIIbβ3 are distinct from those implicated in its interaction to α5β1, suggesting that sCD154 may bind simultaneously to different receptors. Indeed, sCD154 can bind simultaneously to CD40 and α5β1 and biologically activate human monocytic U937 cells expressing both receptors. The simultaneous engagement of CD40 and α5β1 activates the mitogen-activated protein kinases, p38, and extracellular signal-related kinases 1/2 and synergizes in the release of inflammatory mediators MMP-2 and -9, suggesting a cross-talk between these receptors.

Highlights

CD154, an immuno-inflammatory molecule, binds to four receptors
Binding of CD154 to BJAB Cells Is Inhibited by sCD40, whereas Its Binding to U937 Cells Is Inhibited by ␣5␤1 and ␣IIb␤3—To verify the possible concomitant binding of soluble CD154 (sCD154) to two receptors and determine the nature of the interaction of CD154 with various receptors, we first investigated the ability of soluble CD40-Fc, ␣5␤1, or ␣IIb␤3 to modulate the binding of sCD154 to CD40 or ␣5␤1 expressed on the cell surface
Our results indicate that the binding of sCD154-Alexa to BJAB cells was only inhibited by sCD40-Fc (94.6 Ϯ 2.1% inhibition, Fig. 2, A and C)

Summary

Background

CD154, an immuno-inflammatory molecule, binds to four receptors. Results: CD154 differentially binds its various receptors and is capable of simultaneously interacting with multiple ones, inducing synergistic responses in monocytes. SCD154 can bind simultaneously to CD40 and ␣5␤1 and biologically activate human monocytic U937 cells expressing both receptors. Studies using CD154Ϫ/Ϫ and CD40Ϫ/Ϫ mice have suggested that CD154 may bind other receptors [11] In support of this hypothesis, it has been demonstrated that sCD154 interacts with ␣IIb␤3 (GPIIb/IIIa), an integrin expressed on the megakaryocyteplatelet lineage, mast cells, and hematopoietic progenitor cells [12]. This integrin was found to be implicated in atherosclerosis development via its role in leukocyte adhesion to the endothelial layer and their subsequent trans-endothelial migration [18] Taken together, these results clearly indicate that CD154 interacts with four different receptors (CD40, ␣IIb␤3, ␣5␤1, and ␣M␤2) of biological relevance. Simultaneous interaction of CD154 with CD40 and ␣5␤1 on CD40-transfected U937 cells generates a cross-talk between these receptors and induces rapid p38 and ERK1/2 activation, which lead to enhanced MMP-2 and MMP-9 secretion

EXPERIMENTAL PROCEDURES

The abbreviation used is

RESULTS

DISCUSSION