Identification of the Product of Growth Arrest-specific Gene 6 as a Common Ligand for Axl, Sky, and Mer Receptor Tyrosine Kinases

Kyoko Nagata,Kazumasa Ohashi,Toru Nakano,Hitoshi Arita,Chen Zong,Hidesaburo Hanafusa,Kensaku Mizuno

doi:10.1074/jbc.271.47.30022

Abstract

Axl, Sky, and Mer, members of an Axl/Sky receptor tyrosine kinase subfamily, are typified by the cell adhesion molecule-related extracellular domain. The product of growth arrest-specific gene 6 (Gas6), structurally homologous to the anticoagulant protein S, was recently identified as the ligand for Axl and Sky, but the ligand for Mer remained unknown. We have now obtained evidence that Gas6 can also function as a ligand for Mer. Co-precipitation analysis, using soluble receptors of Axl, Sky, and Mer (Axl-Fc, Sky-Fc, and Mer-Fc) composed of the extracellular domain of receptors fused to the Fc domain of immunoglobulin G1, clearly showed that Gas6, but not protein S, specifically bound to Axl-Fc, Sky-Fc, and Mer-Fc fusion proteins. Quantitative kinetic analyses using a BIAcore biosensor instrument revealed dissociation constants (Kd) of the binding of rat Gas6 to Axl-Fc, Sky-Fc, and Mer-Fc are 0.4, 2.7, and 29 nM, respectively. We also found that Gas6 stimulated tyrosine phosphorylation of Axl, Sky, and Mer receptors ectopically expressed in Chinese hamster ovary cells. Taken together, these findings suggest that Gas6 is a common ligand for Axl, Sky, and Mer, all known members of an Axl/Sky receptor subfamily.

Highlights

Axl, Sky, and Mer, members of an Axl/Sky receptor tyrosine kinase subfamily, are typified by the cell adhesion molecule-related extracellular domain
Binding of Gas6 to Axl, Sky, and Mer-Fc Fusion Proteins, as Measured by Co-precipitation Analysis—To characterize and compare in parallel experiments the binding potential of Gas6 and protein S to three distinct members (Axl, Sky, and Mer) of an Axl/Sky receptor subfamily, we prepared chimeric soluble receptors (Axl-Fc, Sky-Fc, and Mer-Fc) composed of the extracellular ligand-binding domain of these receptors fused to the Fc region of the human immunoglobulin IgG1 heavy chain
We prepared Met-Fc, a chimeric protein composed of the extracellular domain of c-Met and the Fc region of IgG1

Summary

Introduction

Sky, and Mer, members of an Axl/Sky receptor tyrosine kinase subfamily, are typified by the cell adhesion molecule-related extracellular domain. We found that Gas6 stimulated tyrosine phosphorylation of Axl, Sky, and Mer receptors ectopically expressed in Chinese hamster ovary cells.

Results

Conclusion