IDENTIFICATION OF OSTEOPONTIN AS A NOVEL LIGAND FOR THE INTEGRIN ALPHA-8-BETA-1 AND POTENTIAL ROLES FOR THIS INTEGRIN-LIGAND INTERACTION INKIDNEY MORPHOGENESIS

Epithelio-mesenchymal interactions during kidney organogenesis are dis rupted in integrin alpha 8 beta 1-deficient mice. However, the known l igands for integrin alpha 8 beta 1-fibronectin, vitronectin, and tenas cin-C-are not appropriately localized to mediate all alpha 8 beta 1 fu nctions in the kidney. Using a method of general utility for determini ng the distribution of unknown Fntegrin ligands in situ and biochemica l characterization of these ligands, we identified osteopontin (OPN) a s a ligand for alpha 8 beta 1. We have coexpressed the extracellular d omains of the mouse alpha 8 and beta 1 integrin subunits as a soluble heterodimer with one subunit fused to alkaline phosphatase (AP) and ha ve used the alpha 8 beta 1-AP chimera as a histochemical reagent on se ctions of mouse embryos. Ligand localization with alpha 8 beta 1-AP in developing bone and kidney was observed to be overlapping with the di stribution of OPN. In ''far Western'' blots of mouse embryonic protein extracts, bands were detected with sizes corresponding to fibronectin , vitronectin, and unknown proteins, one of which was identical to the size of OPN. In a solid-phase binding assay we demonstrated that puri fied OPN binds specifically to alpha 8 beta 1-AP. Cell adhesion assays using K562 cells expressing alpha 8 beta 1 were used to confirm this result. Together with a recent report that anti-OPN antibodies disrupt kidney morphogenesis, our results suggest that interactions between O PN and integrin alpha 8 beta 1 may help regulate kidney development an d other morphogenetic processes.