Affixin Interacts with α-Actinin and Mediates Integrin Signaling for Reorganization of F-Actin Induced by Initial Cell-Substrate Interaction

• Published in: The Journal of cell biology Vol. 165; no. 4; pp. 539 - 551
• Main Authors: Yamaji, Satoshi, Suzuki, Atsushi, Kanamori, Heiwa, Mishima, Wataru, Yoshimi, Ryusuke, Takasaki, Hirotaka, Takabayashi, Maki, Fujimaki, Katsumichi, Fujisawa, Shin, Ohno, Shigeo, Ishigatsubo, Yoshiaki
• Format: Journal Article
• Language: English
• Published: United States Rockefeller University Press 24.05.2004; The Rockefeller University Press
• Subjects: Actinin - genetics; Actinin - metabolism; Actinin - physiology; Actins; Actins - metabolism; Animals; Antibodies; Binding Sites - physiology; Cell adhesion; Cell adhesion & migration; Cell Adhesion - physiology; Cell Movement - physiology; Cell Surface Extensions - metabolism; Cell Surface Extensions - ultrastructure; Cells; Cellular biology; Cellular immunity; CHO Cells; COS Cells; Cricetinae; Cultured cells; Cytoskeleton; Down-Regulation - genetics; HeLa cells; Humans; Integrins; Integrins - metabolism; Microfilament Proteins; Peptides - genetics; Peptides - metabolism; Protein Binding - physiology; Protein Structure, Tertiary - physiology; Protein-Serine-Threonine Kinases - metabolism; Pseudopodia - metabolism; Pseudopodia - ultrastructure; RNA Interference; Signal Transduction - physiology; Signaling; Small interfering RNA; Transfection
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.200308141

The linking of integrin to cytoskeleton is a critical event for an effective cell migration. Previously, we have reported that a novel integrin-linked kinase (ILK)-binding protein, affixin, is closely involved in the linkage between integrin and cytoskeleton in combination with ILK. In the present work, we demonstrated that the second calponin homology domain of affixin directly interacts with α-actinin in an ILK kinase activity-dependent manner, suggesting that integrin-ILK signaling evoked by substrate adhesion induces affixin-α-actinin interaction. The overexpression of a peptide corresponding to the α-actinin-binding site of affixin as well as the knockdown of endogenous affixin by small interference RNA resulted in the blockade of cell spreading. Time-lapse observation revealed that in both experiments cells were round with small peripheral blebs and failed to develop lamellipodia, suggesting that the ILK-affixin complex serves as an integrin-anchoring site for α-actinin and thereby mediates integrin signaling to α-actinin, which has been shown to play a critical role in actin polymerization at focal adhesions.