ADAM 12, a member of the ADAM family of transmembrane metalloprotease-disintegrins, has been implicated previously in the differentiation of skeletal myoblasts. In the present study, we show that the cytoplasmic tail of mouse ADAM 12 interacts in vitro and in vivo with the Src homology 3 domain of the p85␣ regulatory subunit of phosphatidylinositol (PI) 3-kinase. By site-directed mutagenesis, we have identified three p85␣-binding sites in ADAM 12 involving PXXP motifs located at amino acids

more »

... 828, 833-836, and 884 -887. Using green fluorescent protein (GFP)-pleckstrin homology (PH) domain fusion protein as a probe for PI 3-kinase lipid products, we have further demonstrated that expression of ADAM 12 in C2C12 cells resulted in translocation of GFP-PH to the plasma membrane. This suggests that transmembrane ADAM 12, by providing docking sites for the Src homology 3 domain of p85␣, activates PI 3-kinase by mediating its recruitment to the membrane. Because PI 3-kinase is critical for terminal differentiation of myoblasts, and because expression of ADAM 12 is up-regulated at the onset of the differentiation process, ADAM 12-mediated activation may constitute one of the regulatory mechanisms for PI 3-kinase during myoblast differentiation. . 1 The abbreviations used are: ADAM, protein containing a disintegrin and metalloprotease; aa, amino acid; ARNO, Arf nucleotide binding site opener; SH3, Src homology 3 domain; SH2, Src homology 2 domain;