Regulation of cell adhesion to collagen via [beta]1 integrins is dependent on interactions of filamin A with vimentin and protein kinase C epsilon

• Published in: Experimental cell research 2010-07, Vol.316 (11), p.1829
• Main Authors: Kim, Hugh, Nakamura, Fumihiko, Lee, Wilson, Hong, Claire, Pérez-Sala, Dolores, McCulloch, Christopher A
• Format: Article
• Language: English
• Subjects: Cell adhesion & migration; Cellular biology; Kinases; Mutation; Proteins
• ISSN: 0014-4827; 1090-2422
• DOI: 10.1016/j.yexcr.2010.02.007

Cell adhesion and spreading on collagen, which are essential processes for development and wound healing in mammals, are mediated by [beta]1 integrins and the actin and intermediate filament cytoskeletons. The mechanisms by which these separate cytoskeletal systems interact to regulate [beta]1 integrins and cell spreading are poorly defined. We previously reported that the actin cross-linking protein filamin A binds the intermediate filament protein vimentin and that these two proteins co-regulate cell spreading. Here we used deletional mutants of filamin A to define filamin A-vimentin interactions and the subsequent phosphorylation and re-distribution of vimentin during cell spreading on collagen. Imaging of fixed and live cell preparations showed that phosphorylated vimentin is translocated to the cell membrane during spreading. Knockdown of filamin A inhibited cell spreading and the phosphorylation and re-distribution of vimentin. Knockdown of filamin A and/or vimentin reduced the cell surface expression and activation of [beta]1 integrins, as indicated by immunoblotting of plasma membrane-associated proteins and shear force assays. In vitro pull-down assays using filamin A mutants showed that both vimentin and protein kinase C bind to repeats 1-8 of filamin A. Reconstitution of filamin-A-deficient cells with full-length filamin A or filamin A repeats 1-8 restored cell spreading, vimentin phosphorylation, and the cell surface expression of [beta]1 integrins. We conclude that the binding of filamin A to vimentin and protein kinase C[straight epsilon] is an essential regulatory step for the trafficking and activation of [beta]1 integrins and cell spreading on collagen. [PUBLICATION ABSTRACT]