Spreading dynamics of biomimetic actin cortices

Abstract

Reconstituted systems mimicking cells are interesting tools for understanding the details of cell behavior. Here, we use an experimental system that mimics cellular actin cortices, namely liposomes developing an actin shell close to their inner membrane, and we study their dynamics of spreading. We show that depending on the morphology of the actin shell inside the liposome, spreading dynamics is either reminiscent of a bare liposome (in the case of a sparse actin shell) or of a cell (in the case of a continuous actin shell). We use a mechanical model that qualitatively accounts for the shape of the experimental curves. From the data on spreading dynamics, we extract characteristic times that are consistent with mechanical estimates. The mechanical characterization of such stripped-down experimental systems paves the way for a more complex design closer to a cell. We report here the first step in building an artificial cell and studying its mechanics.

Figure 1

Dynamics of spreading. Area of contact as a function of time observed by RICM (A) for low χ and (B) for high χ conditions. (Insets) Time lapse of RICM outline of spreading area with indicated time interval; scale bars are 5 μm. Squares in panel A are curves for bare liposomes, solid dots are curves for different values of χ, as indicated in the insets.

Figure 2

Actin cortices observed by confocal microscopy in a small liposome (radius R = 4.5 μm, left column) and in a large liposome (radius R = 18 μm, right column). (A and B) Three-dimensional top view. (C and D) Equatorial fluorescence as a function of time. (E and F) Three-dimensional bottom view, with the plane of contact indicated by the four solid corners; the open lines indicate the contour of the contact area. The small liposome (left column) appears patchier than the large one (right column). Unit square = 5 μm. (G and I) Fluorescence images of the contact area observed by epifluorescence microscopy for different value of χ. (H and J) Calculations of variance, χ in the fluorescence intensity in panels G and I, over a window of two pixels. The χ parameter characterizes the density of actin.

Figure 3

Tracking of actin patches distinguishes continuous from discontinuous cortices. (A) Fluorescent actin patches within an apical confocal slice. The actin cortex may be discontinuous with low χ (top left, the slice is taken at 31 μm from the surface), or continuous with high χ-cortices (bottom left, the slice is taken at 17 μm from the surface). Isosurface rendering and surface tracking of the actin patches maps out their trajectories over time (A, right). Shaded regions are the tracked actin patches, and the trajectories are colored to reflect time, getting darker as time progresses. Grid is 1 μm. (B) MSD for the individual surfaces in panel A, on a linear plot, and log plot (inset). (Lines) Linear fits of the MSD.

Figure 4

Liposome at steady state of spreading. (A) Schematic representation of an adhered liposome at steady state with notations. Surface tension (or force per unit contact length) are γ_SW between the solid substrate and the aqueous solution surrounding the liposome, γ_SL between the substrate/liposome surface, and σ_f is the final membrane tension. (B) The adhered liposome is observed by confocal microscopy and characterized by the contact angle θ^∗ between the liposome and the coated surface. Unit square is 1 μm. (C) Contact angle θ^∗ distribution for liposomes of all χ. The plot is normalized by its maximum. The average value is 〈θ^∗〉 = 48.8°. The histogram is fit by a Gaussian (line) with a variance of 252.3.

Figure 5

Spreading dynamics of liposomes with varying actin content. Spreading of (A) bare, (B) low χ, and (C) high χ liposomes on polyhistidine-coated coverslips. The contact area is normalized by the steady-state spread area A^∗, and the time is normalized by the fitted time constant for each curve, τ_m for panels A and B, and τ_c for panel C. (D) Fitted time constants τ_m for bare liposomes (open circles) and liposomes displaying a low χ (squares), with τ_c for liposomes displaying a high χ (solid circles) as a function of the χ-value. Bare liposomes correspond to χ = 0. (E) Schematic representation of an adhered liposome during early spreading with notations.