Shedding of Kidney Injury Molecule-1, a Putative Adhesion Protein Involved in Renal Regeneration

Abstract

KIM-1 (kidney injury molecule-1) is a type I transmembrane glycoprotein expressed on dedifferentiated renal proximal tubule epithelial cells undergoing regeneration after toxic or ischemic injury. The extracellular domain of KIM-1 is composed of an immunoglobulin-like domain topping a long mucin-like domain, a structure that points to a possible role in cell adhesion by homology to several known adhesion proteins. Two splice variants (a and b), of the human KIM-1 having identical extracellular domains, differ in their cytoplasmic domains and tissue distributions. In this study, we report that the KIM-1b transcript is expressed predominantly in adult human kidney. We describe the generation of 10 monoclonal antibodies against the extracellular domain of human KIM-1, the mapping of their binding sites, and their use in identifying various forms of the protein. We show that human KIM-1b is expressed in adult kidney cell lines, and we demonstrate that a soluble form of KIM-1 is shed constitutively into the culture medium of the cell lines expressing endogenous or recombinant KIM-1b by membrane-proximal cleavage. A monoclonal antibody that binds at or close to the proteolytic site can partially block the shedding of KIM-1. Release of soluble KIM-1 is enhanced by activating the cells with phorbol 12-myristate 13-acetate and can be inhibited with two metalloproteinase inhibitors, BB-94 (Batimastat) and GM6001 (Ilomastat), suggesting that the cleavage is mediated by a metalloproteinase. We propose that the shedding of KIM-1 in the kidney undergoing regeneration constitutes an active mechanism allowing dedifferentiated regenerating cells to scatter on denuded patches of the basement membrane and reconstitute a continuous epithelial layer.