Abstract
BackgroundThe cytosol of amoeba cells controls the membrane deformation during their motion in vivo. To investigate such ability of the cytosol of amoeba cell, Dictyostelium discoideum (Dictyostelium), in vitro, we used lipids extracted from Dictyostelium and commercially available phospholipids, and prepared substrate-supported lipid membrane patches on the micrometer scale by spin coating.ResultsWe found that the spin coater holder, which has pores (pore size = 3.1 mm) of negative pressure to hold the cover glass induced the concave surface of the cover glass. The membrane lipid patches were formed at each position in the vicinity of the holder pores and their sizes were in the range of 2.7 to 3.2 × 104 μm2. After addition of the cytosol extracted from Dictyostelium to the lipid membrane patches, through time-lapse observation with a confocal laser scanning fluorescence microscope, we observed an autonomous buckling of the Dictyostelium lipid patches and localized behaviours of proteins found within.ConclusionThe current method serves as the novel technique for the preparation of film patches in which the positions of patches are controlled by the holder pores without fabricating, modifying, and arranging the chemical properties of the solution components of lipids. The findings imply that lipid-binding proteins in the cytosol were adsorbed and accumulated within the Dictyostelium lipid patches, inducing the transformation of the cell-sized patch.Electronic supplementary materialThe online version of this article (doi:10.1186/1754-1611-9-3) contains supplementary material, which is available to authorized users.
Highlights
The cytosol of amoeba cells controls the membrane deformation during their motion in vivo
Micrometer-sized patches of lipid membrane film prepared by spin coating The spin coating method for the lipid membrane film supported on a glass substrate conventionally enables us to prepare heterogeneous films using a lipid mixture [10]
After extraction of the lipid mixture from the wild-type Dictyostelium, AX4 cells, we observed the lipid membrane film of Dictyostelium prepared by the spin coating method on the millimeter scale by light microscopy
Summary
The cytosol of amoeba cells controls the membrane deformation during their motion in vivo To investigate such ability of the cytosol of amoeba cell, Dictyostelium discoideum (Dictyostelium), in vitro, we used lipids extracted from Dictyostelium and commercially available phospholipids, and prepared substrate-supported lipid membrane patches on the micrometer scale by spin coating. Model construction for cell dynamics, such as membrane buckling [4,5], has attracted the attention of researchers because such cell dynamics are closely related to amoeba motion which highly functionalized with the coupling of cellular should be completely purified and mixed in a sequential manner We overcame this obstacle by using lipids and cytosol extracted from an amoeba cell and constructing micrometer-sized substrate-contacting lipidprotein membrane patches. In order to trace the transformation of such lipid membrane film in a three-dimensional manner after the addition of the cytosolic extract, we performed time lapse observations using a confocal laser scanning fluorescence microscope
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