Role of Focal Adhesion Kinase Ser-732 Phosphorylation in Centrosome Function during Mitosis

Ann Y.J Park,Tang-Long Shen,Shu Chien,Jun-Lin Guan

doi:10.1074/jbc.m809040200

Abstract

Focal adhesion kinase (FAK) is the major cytoplasmic tyrosine kinase in focal adhesions and a critical mediator of integrin signaling in a variety of cells, including endothelial cells (ECs). Here we describe a new function for FAK in the regulation of centrosome functions in a Ser-732 phosphorylation-dependent manner during mitosis. Deletion of FAK in primary ECs causes increases in centrosome numbers, multipolar and disorganized spindles, and unaligned chromosomes during mitosis. Re-expression of wild-type FAK, but not S732A mutant, rescued these mitotic defects, suggesting a role for Ser-732 phosphorylation in the regulation of centrosomal functions. Consistent with this possibility, Ser-732-phosphorylated FAK was found to co-localize in centrosomes in mitotic cells. FAK also associated with cytoplasmic dynein in a Ser-732 phosphorylation-dependent manner. Further analysis in FAK-null primary ECs showed that S732A mutant could rescue EC migration but not proliferation or tubulogenesis in vitro. Last, we showed that deletion of FAK in ECs reduced tumor angiogenesis in vivo, which could be restored by re-expression of wild-type FAK but not S732A mutant. Together, these studies demonstrated a novel role for Ser-732 phosphorylation of FAK in the regulation of centrosome function during mitosis, which may contribute to EC proliferation and angiogenesis.

Highlights

Focal adhesion kinase (FAK), including residues Tyr-576 and Tyr-577 in its activation loop, to further activate FAK and Tyr-925 to create binding sites for the adaptor molecule Grb2 [7,8,9]
In this study we present data showing a novel function of FAK in the regulation of centrosomal functions in a Ser-732 phosphorylation-dependent manner in endothelial cells (ECs) during mitosis, which plays a role in the regulation of EC proliferation and tubulogenesis in vitro and tumor angiogenesis in vivo
Primary ECs were isolated from floxed FAK mice and were infected by a recombinant adenovirus encoding Cre recombinase (Ad-Cre) to delete endogenous FAK or by a control recombinant adenovirus encoding lacZ to produce FAKϪ/Ϫ ECs or control FAKϩ/ϩ ECs, respectively, as described previously [27]

Summary

Introduction

FAK, including residues Tyr-576 and Tyr-577 in its activation loop, to further activate FAK and Tyr-925 to create binding sites for the adaptor molecule Grb2 [7,8,9]. In this study we present data showing a novel function of FAK in the regulation of centrosomal functions in a Ser-732 phosphorylation-dependent manner in ECs during mitosis, which plays a role in the regulation of EC proliferation and tubulogenesis in vitro and tumor angiogenesis in vivo.

Results

Conclusion