Persistence Length of Fascin-cross-linked Actin Filament Bundles in Solution and the in Vitro Motility Assay

Background: Bundles of unipolar actin filaments (F-actin), cross-linked via the actin-binding protein fascin, are important in filopodia of motile cells and stereocilia of inner ear sensory cells. However, such bundles are also useful as shuttles in myosin-driven nanotechnological applications. Therefore, and for elucidating aspects of biological function, we investigate if the bundle tendency to follow straight paths (quantified by path persistence length) when propelled by myosin motors is directly determined by material properties quantified by persistence length of thermally fluctuating bundles.

Methods: Fluorescent bundles, labeled with rhodamine-phalloidin, were studied at fascin:actin molar ratios: 0:1 (F-actin), 1:7, 1:4 and 1:2. Persistence lengths (Lp) were obtained by fitting the cosine correlation function (CCF) to a single exponential function: <cos(θ(0)-θ(s))>=exp(-s/(2Lp)) where θ(s) is tangent angle; s: path or contour lengths. < > denotes averaging over filaments.

Results: Bundle-Lp (bundles<15μm long) increased from ~10 to 150μm with increased fascin:actin ratio. The increase was similar for path-Lp (path<15μm), with highly linear correlation. For longer bundle paths, the CCF-decay deviated from a single exponential, consistent with superimposition of the random path with a circular path as suggested by theoretical analysis.

Conclusions: Fascin-actin bundles have similar path-Lp and bundle-Lp, both increasing with fascin:actin ratio. Path-Lp is determined by the flexural rigidity of the bundle.

General Significance: The findings give general insight into mechanics of cytoskeletal polymers that interact with molecular motors, aid rational development of nanotechnological applications and have implications for structure and in vivo functions of fascin-actin bundles.