Abstract 226: the Focal Adhesion Kinase is Phosphorylated on Serine732 by a Growth Factor-Dependent Pathway and Contributes to the Proliferation of Tumor Cells

Abstract Focal adhesion kinase (FAK) is a tyrosine kinase localized at the site of focal adhesions (FA). FAK is identified as a key mediator of signaling by integrins, the major family of cell surface receptors for extracellular matrix, as well as by other receptors in both normal and cancer cells. FAK plays a prominent role in tumor progression and metastasis regulating cellular processes including migration, invasion, epithelial to mesenchymal transition, and angiogenesis. Integrin-FAK signaling has been shown to activate a number of biological mechanisms through phosphorylation and protein-protein interactions promoting tumorigenesis. FAK seems also to have a role in the proliferation of tumor cells by its biochemical and biological association to cytoplasmic kinases such as src and ERK. However, the mechanisms that involve FAK in those processes are not yet clarify. The best characterized FAK phosporylation event is the auto-phosphorylation at the tyrosine397 (Tyr397) which creates a motif that is recognized by various SH2 domain-containing proteins, such as src which phosphorylates FAK on Tyr576. The c-terminal domain of FAK undergoes to several serine-phosphorylation events whose role is not well-understood. Previous studies in neural and endothelial cells (EC) have shown a new role for FAK in the regulation of centrosome functions during mitosis dependent on Ser732 phosphorylation. We have found that EGF stimulation of selected starved melanoma, thyroid and high stage ovarian tumor cells induced, in dividing cells, accumulation of phosphorylated (P-)FAK on Ser732. Inhibition of cell/substrate adhesion with specific anti-integrin antibodies demonstrated that P-FAKSer732 activation is not induced by integrin clustering. Confocal immunofluorescence and immunoprecipitation showed that, in these cells, P-FAKSer732 is localized in a perinuclear site independently from FA and P-FAKTyr397. Treatment of an high proliferative melanoma cell line with the Rock inhibitor Y27632 partially decreased the levels of P-FAKSer732, the cellular proliferation and impaired the mitotic spindle formation. FACS analysis showed that the P-FAKSer732 decreased upon treatment with the MEK inhibitor UO126 in a dose-dependent manner together with the levels of P(Ser 10) -Histone H3 thus demonstrating that FAK is involved in the mitotic process regulated by MEK/ERK activation. Indeed, P-FAKSer732 co-localized with microtubules of the mitotic spindle in proliferating cells and immunoprecipitated with the motor protein dynein and with the acetylated tubulin of polymerized microtubules. The finding of a novel role for FAK in tumor cells might provide the underpinning for therapeutic strategies in selected solid tumors. Partially supported by Italian Ministery of Health and Associazione Italiana per la Ricerca sul Cancro (AIRC). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 226. doi:1538-7445.AM2012-226