Meltrin α cytoplasmic domain interacts with SH3 domains of Src and Grb2 and is phosphorylated by v-Src

Abstract

Meltrin α/ADAM12 is a member of the ADAM/MDC family proteins characterized by the presence of metalloprotease and disintegrin domains. This protein also contains a single transmembrane domain and a relatively long cytoplasmic domain containing several proline-rich sequences. These sequences are compatible with the consensus sequences for binding the Src homology 3 (SH3) domains. To determine whether the proline-rich sequences interact with SH3 domains in several proteins, binding of recombinant SH3 domains to the meltrin α cytoplasmic domain was analysed by pulldown assays. The SH3 domains of Src and Yes bound strongly, but that of Abl or phosphatidylinositol 3-kinase p85 subunit did not. Full-length Grb2/Ash bound strongly, whereas its N-terminal SH3 domain alone did less strongly. Src and Grb2 in bovine brain extracts also bound to meltrin α cytoplasmic domain on affinity resin. Furthermore, immunoprecipitation with a monoclonal antibody to meltrin α resulted in coprecipitation of Src and Grb2 with meltrin α in cell extracts, suggesting that Src and Grb2 are associated in vivo with meltrin α cytoplasmic domain. This notion was also supported by the findings that exogenously expressed meltrin α cytoplasmic domain coexisted with Src and Grb2 on the membrane ruffles. The C-terminal Tyr901 of meltrin α was phosphorylated both in vitro and in cultured cells by v-Src. These results may imply that meltrin α 8cytoplasmic domain is involved in a signal transduction for some biological function through the interaction with SH3-containing proteins.