Regulated association of microtubule-associated protein 2 (MAP2) with Src and Grb2: Evidence for MAP2 as a scaffolding protein

Microtubule-associated protein 2 (MAP2) and tan, which is involved in Alzhe imer's disease, are major cytoskeletal proteins in neurons. These proteins are involved in microtubule assembly and stability, To further characterize MAP2, we took a strategy of identifying potential MAP2 binding partners. T he low molecular weight MAP2c protein has 11 PXXP motifs that are conserved across species, and these PXXP motifs could be potential ligands for Src h omology 3 (SH3) domains. We tested for MAP2 interaction with SH3 domain-con taining proteins. All neuronal MAP2 isoforms bound specifically to the SH3 domains of c-Src and Grb2 in an in vitro glutathione S-transferase-SH3 pull -down assay. Interactions between endogenous proteins were confirmed by co- immunoprecipitation using brain lysate, All three proteins were also found co-expressed in neuronal cell bodies and dendrites. Surprisingly, the SH3 d omain-binding site was mapped to the microtubule-binding domain that contai ns no PXXP motif. Src bound primarily the soluble, non-microtubule-associat ed MAP2c in vitro. This specific MAP2/SH3 domain interaction was inhibited by phosphorylation of MAP2c by the mitogen-activated protein kinase extrace llular signal-regulated kinase 2 but not by protein kinase A. This phosphor ylation-regulated association of MAP2 with proteins of intracellular signal transduction pathways suggests a possible link between cellular signaling and neuronal cytoskeleton, with MAP2 perhaps acting as a molecular scaffold upon which cytoskeleton-modifying proteins assemble and dissociate in resp onse to neuronal activity.