Novel Structures for α-Actinin:F-Actin Interactions and their Implications for Actin–Membrane Attachment and Tension Sensing in the Cytoskeleton

Abstract

We have applied correspondence analysis to electron micrographs of 2-D rafts of F-actin cross-linked with α-actinin on a lipid monolayer to investigate α-actinin:F-actin binding and cross-linking. More than 8000 actin crossover repeats, each with one to five α-actinin molecules bound, were selected, aligned, and grouped to produce class averages of α-actinin cross-links with ∼9-fold improvement in the stochastic signal-to-noise ratio. Measurements and comparative molecular models show variation in the distance separating actin-binding domains and the angle of the α-actinin cross-links. Rafts of F-actin and α-actinin formed predominantly polar 2-D arrays of actin filaments, with occasional insertion of filaments of opposite polarity. Unique to this study are the numbers of α-actinin molecules bound to successive crossovers on the same actin filament. These “monofilament”-bound α-actinin molecules may reflect a new mode of interaction for α-actinin, particularly in protein-dense actin–membrane attachments in focal adhesions. These results suggest that α-actinin is not simply a rigid spacer between actin filaments, but rather a flexible cross-linking, scaffolding, and anchoring protein. We suggest these properties of α-actinin may contribute to tension sensing in actin bundles.