Search for a Novel Gla-containing Growth Factor as the Ligand for Mer Receptor Tyrosine Kinase

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We previously identified that Gas6 is a common ligand for Axi, Sky, and Mer receptor tyrosine kinases. Quantitative binding analysis revealed that the binding affinity of Gas6 to Mer is relatively low, with a Kd value of 29.0 nM.Thus, we assumed the existence of the Gas6-related protein as a ligand for Mer receptor. In this study, we searched for the Gas6-related protein that may function as the preferable ligand for Mer. Gas6 has a structure similar to that of protein S and is composed of a Gla domain, four EGF-like domains and a C-terminal sex hormone-binding globulin (SHBG)- like domain. When examining the role of each domain in receptor-binding and biological activities of Gas6, we found that receptor-binding and mitogenic activities were markedly reduced by inhibiting gamma -carboxylation of the Gla domain, while a Gas6 mutant composed of only an SHBG-like domain retained both of these activities. Thus, the SHBG-like domain is apparently an entity indispensable for Gas6 activities, and gamma -carboxylation of the Gla domain has a regulatory role in retaining the activity of native Gas6. We searched for the cDNA clones coding for Gas6-related proteins, using PCR and low stringency hybridization techniques, but we failed to clone coding, for such protein. Androgen-binding protein, which has low sequence similarity to the SHBG-like domain of Gas6, showed no binding ability to Mer, Sky or Axl receptor. Thus, we have not detected any Gas6-related protein other than protein S.It could be that Gas6-induced activation of Mer require co-factors or co-receptors in living organisms. We also showed that Axl-Fc, a soluble form of AxI receptor composed of the extracellular domain of AxI and the Fc region of immunoglobulin, had an inhibitory function for the growth potentiating activity of Gas6 on vascular smooth muscle cells.