Mechanisms of Signal Transduction
Phosphoinositide Binding Inhibits α-Actinin Bundling Activity*

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α-Actinin is an abundant actin-bundling and adhesion protein that directly links actin filaments to integrin receptors. Previously, in platelet-derived growth factor-treated fibroblasts, we demonstrated that phosphoinositides bind to α-actinin, regulating its localization (Greenwood, J. A., Theibert, A. B., Prestwich, G. D., and Murphy-Ullrich, J. E. (2000) J. Cell Biol. 150, 627– 642). In this study, phosphoinositide binding and regulation of α-actinin function is further characterized. Phosphoinositide binding specificity, determined using a protein-lipid overlay procedure, suggests that α-actinin interacts with phosphates on the 4th and 5th position of the inositol head group. Binding assays and mutational analyses demonstrate that phosphoinositides bind to the calponin homology domain 2 of α-actinin. Phosphoinositide binding inhibited the bundling activity of α-actinin by blocking the interaction of the actin-binding domain with actin filaments. Consistent with these results, excessive bundling of actin filaments was observed in fibroblasts expressing an α-actinin mutant with decreased phosphoinositide affinity. We conclude that the interaction of α-actinin with phosphoinositides regulates actin stress fibers in the cell by controlling the extent to which microfilaments are bundled.

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This work was supported in part by Grant GM 63711 from the National Institutes of Health (to J. A. G.), American Cancer Society Oregon Chapter, Medical Research Foundation of Oregon, and Northwest Health Foundation. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Supported by summer undergraduate research fellowships funded by Howard Hughes Medical Institute Grant 57003741 (to Oregon State University).