THE RAS TARGET AF-6 IS A SUBSTRATE OF THE FAM DEUBIQUITINATING ENZYME

The Ras target AF-6 has been shown to serve as one of the peripheral c omponents of cell-cell adhesions, and is thought to participate in cel l-cell adhesion regulation downstream of Ras. We here purified an AF-6 -interacting protein with a molecular mass of similar to 220 kD (p220) to investigate the function of AF-6 at cell-cell adhesions. The pepti de sequences of p220 were identical to the amino acid sequences of mou se Fam. Fam is homologous to a deubiquitinating enzyme in Drosophila, the product of the fat facets gene. Recent genetic analyses indicate t hat the deubiquitinating activity of the fnr facets product plays a cr itical role in controlling the cell fate. We found that Fam accumulate d at the cell-cell contact sites of MDCKII cells, but not at free ends of plasma membranes. Fam was partially colocalized with AF-6 and inte racted with AF-6 in vivo and in vitro. We also showed that AF-6 was ub iquitinated in intact cells, and that Fam prevented the ubiquitination of AF-6. These results indicate that AF-6 forms a complex with and se rves as one of the substrates for Fam, and suggest that the degradatio n of peripheral components of cell-cell adhesions may be regulated by Fam.