JAM4, a Junctional Cell Adhesion Molecule Interacting with a Tight Junction Protein, MAGI-1

Abstract

MAGI-1 is a membrane-associated guanylate kinase protein at tight junctions in epithelial cells. It interacts with various molecules and functions as a scaffold protein at cell junctions. We report here a novel MAGI-1-binding protein that we named junctional adhesion molecule 4 (JAM4). JAM4 belongs to an immunoglobulin protein family. JAM4 was colocalized with ZO-1 in kidney glomeruli and in intestinal epithelial cells. Biochemical in vitro studies revealed that JAM4 bound to MAGI-1 but not to ZO-1, whereas JAM1 did not bind to MAGI-1. JAM4 and MAGI-1 interacted with each other and formed clusters in COS-7 cells when coexpressed. JAM4 mediated calcium-independent homophilic adhesion and was accumulated at cell-cell contacts when expressed in L cells. MAGI-1, ZO-1, and occludin were recruited to JAM4-based cell contacts. JAM4 also reduced the permeability of CHO cell monolayers. MAGI-1 strengthened JAM4-mediated cell adhesion in L cells and sealing effects in CHO cells. These findings suggest that JAM4 together with MAGI-1 provides an adhesion machinery at tight junctions, which may regulate the permeability of kidney glomerulus and small intestinal epithelial cells.

ACKNOWLEDGMENTS

This study was supported by grants-in-aids for Scientific Research and Special Coordination Funds for Promoting Science and Technology from the Ministry of Education, Culture, Sports, Science, and Technology and a grant from Yamanouchi Foundation for Research on Metabolic Disorders (2000).

We thank S. Tsukita (Kyoto University) for cDNA of mouse ZO-1, rat anti-occludin antibody, and MDCKII cells, M. Takeichi for MTD-1A cells, Y. Nakamura (University of Tokyo) for cDNA of human MAGI-1, S. Uchida and S. Sasaki (Tokyo Medical and Dental University) for LLC-PK1 cells, T. Kitamura (University of Tokyo) for pMXpuro vector, G. Noran (Stanford University) for Phoenix ampho cells, S. Hollenberg (Oregon Health Sciences University) for reagents for yeast two-hybrid screening, and T.-C. He and B. Vogelstein (Johns Hopkins Oncology Center) for the pAdEasy system. We thank N. Terada, S. Ichinose, and K. Terashima (Tokyo Medical and Dental University) for valuable advice. We also thank C. Rokukawa and M. Miyahara-Tenkatsu for skillful technical assistance.