Cortactin phosphorylation sites mapped by mass spectrometry.

Cortactin is a scaffolding protein that is targeted to sites of actin polymerization, including lamellipodia in migrating cells, cell-cell junctions in epithelial cells, growth cones of neurons, podosomes of osteoclasts, invadopodia of tumor cells, and sites of actin rearrangement induced by pathogenic bacteria and viruses (Daly, 2004; Lua and Low, 2005; Pfaff and Jurdic, 2001). Although it is phosphorylated in response to a number of stimuli including growth factors, bacterial invasion, platelet activation, transformation and integrin-mediated adhesion, the only sites of phosphorylation that have been characterized to date are Y421, Y466 and Y482 (Src sites that regulate cell migration) and S405 and S418 (ERK sites that regulate N-WASP-mediated actin polymerization) (Campbell et al., 1999; Huang et al., 1998; MartinezQuiles et al., 2004; Weed and Parsons, 2001). To better understand how the phosphorylation of cortactin contributes to the dynamic regulation of actin structures, we mapped the repertoire of phosphorylation sites within cortactin by mass spectrometry (MS). Herein, we report the identification of 17 new sites of phosphorylation (12 serine, four threonine and one tyrosine residues). The distribution of phosphorylation sites on well-conserved residues within cortactin supports a role for these modifications in the regulation of cortactin function.