Analysis of the shape fluctuations of reconstituted membranes using GUVs made from lipid extracts of invertebrates

Abstract

Changes in the physical properties of the lipid matrix of cell membranes have repeatedly been proposed to underlie stresses associated with e.g. drought, cold and xenobiotics. Therefore, the ability to experimentally monitor such properties is central to the fundamental physiological understanding of adaptive changes. Here, we test the analysis of shape fluctuations in membranes composed of lipid extracts from two soil invertebrates, and show that theories and experimental approaches previously developed for simpler liposomes may be applied directly to reconstituted membrane lipids. Specifically, we show how the bending rigidity of giant unilamellar liposomes of lipid extracts can be determined precisely. We suggest that future measurements of this parameter could elucidate mechanisms of adaptive processes such as changes in lipid composition and accumulation of protective osmolytes.

Keywords: Cold tolerance; Enchytraeus albidus; Folsomia candida; Membrane fluidity.

Fig. 1.. Bunches of GUVs obtained from lipid extracts of Folsomia candida springtails visualized by phase contrast microscopy.

The electrode appears in black at the top part of the picture, while the giant vesicles in the focal plane are visible as grey circles. Scale bar: 10 µm.

Fig. 2.. Illustrations of the fluctuation characteristics of a GUV (radius 19.8 µm) made from Enchytraeus albidus membrane lipid extracts.

(a) Spectrum of the amplitudes of decomposition of the angular autocorrelation function in the Legendre polynomial basis for the mode 5. (b) Experimental distribution for the Fourier amplitudes of the contour fluctuations, α⁵ or β⁵, that can be parameterized by a Gaussian fit represented in red line. The mode 5 has been chosen randomly among the mid-range modes (4–10), which have good statistics and are not limited by the video integration time.

Fig. 3.. Illustrations of the fitting procedure for a GUV made from Enchytraeus albidus membrane lipid extracts of radius equal to 19.8 µm.

(a) Experimental distributions for the amplitudes of decomposition of the angular autocorrelation function in the cosine basis, ξ⁵, that can be parameterized by an exponential fit represented in red line. (b) Dependence of Δⁿ as a function of the mode number (close dots) up to the 11th mode and the resulting fit in full line giving κ = 24.48 k_BT and  = 28.85.

Fig. 4.

Individual bending rigidity measurements of POPC GUVs (•), and reconstituted GUVs made from membrane lipid extracts of Folsomia candida (▪) and Enchytraeus albidus (▴), represented in close symbols, while the resulting averages with the standard deviations are shown with empty symbols.

Fig. 5.

(a) Normalized temporal correlation function for mode 5 for a GUV made from Enchytraeus albidus membrane lipid extracts, whose decay is exponentially fitted at early times (solid line). The corresponding decay constants (relaxation times) for the exponentials are depicted in (b) as a function of mode number and scale as τ∼n^−1.67 (solid line).