The S100 Family Heterodimer, MRP-8/14, Binds with High Affinity to Heparin and Heparan Sulfate Glycosaminoglycans on Endothelial Cells

Matthew J Robinson,Philippe Tessier,Richard Poulsom,Nancy Hogg

doi:10.1074/jbc.m102950200

Abstract

The S100 family proteins MRP-8 (S100A8) and MRP-14 (S100A9) form a heterodimer that is abundantly expressed in neutrophils, monocytes, and some secretory epithelia. In inflamed tissues, the MRP-8/14 complex is deposited onto the endothelium of venules associated with extravasating leukocytes. To explore the receptor interactions of MRP-8/14, we use a model system in which the purified MRP-8/14 complex binds to the cell surface of an endothelial cell line, HMEC-1. This interaction is mediated by the MRP-14 subunit and is mirrored by recombinant MRP-14 alone. The cell surface binding of MRP-14 was blocked by heparin, heparan sulfate, and chondroitin sulfate B, and the binding sites were sensitive to heparinase I and trypsin treatment but not to chondroitinase ABC. Furthermore MRP-8/14 and MRP-14 did not bind to a glycosaminoglycan-minus cell line. MRP-14 has a high affinity for heparin (K(d) = 6.1 +/- 3.4 nm), and this interaction mimicked that with the endothelial cells. We therefore conclude that the MRP-8/14 complex binds to endothelial cells via the MRP-14 subunit interacting chiefly with heparan sulfate proteoglycans. CD36 and RAGE, two other putative receptors for MRP-8/14, were not expressed by HMEC-1 cells. This binding activity may explain the immobilization of the MRP-8/14 complex on endothelium that is observed in vivo.

Highlights

The S100 proteins are a family of small (10 –14 kDa) calciumbinding proteins [1, 2]
The CD45-positive total leukocyte infiltrate (Fig. 1B) contained CD15-positive neutrophils and CD68-positive monocytes but was negative for MRP-8 and -14 (Fig. 1A). Together these results suggest that in an inflammatory setting, myeloid cells lose expression of MRP-8 and -14 during transmigration, and this can be associated with MRP positive endothelium
We show for the first time that MRP-8/14-positive vessels adjacent to an inflammatory site do not synthesize the MRP proteins but bind MRP-8/14 that appears to have been released by transmigrating myeloid cells

Summary

Introduction

The S100 proteins are a family of small (10 –14 kDa) calciumbinding proteins [1, 2]. The majority of the S100 genes are tightly clustered together on chromosome 1q21 in man and chromosome 3 in the mouse, but the individual proteins are expressed in distinctive cell types. To explore the receptor interactions of MRP-8/14, we use a model system in which the purified MRP-8/14 complex binds to the cell surface of an endothelial cell line, HMEC-1. As determined by immunohistochemistry and enzyme-linked immunosorbent assay for the MRP-8/-14 complex, the HMEC-1 cells were found not to express the MRP proteins even after stimulation (data not shown).

Results

Conclusion