Fig. 1: EV membrane physical properties and protein content mirror those of lipid rafts. | Nature Communications

Fig. 1: EV membrane physical properties and protein content mirror those of lipid rafts.

From: Enhancing extracellular vesicle cargo loading and functional delivery by engineering protein-lipid interactions

Fig. 1

a Schematic of our hypothesis that lipid raft association could be used as a handle to load proteins into EVs. b Overall analysis and experimental workflow. We used RaftProt 2.0 and Exocarta to understand features of proteins found in EVs and lipid rafts (I); we then built a library of structurally diverse proteins to understand how such features affect protein trafficking, interactions with lipid rafts, and loading into EVs (II); applying these design rules, we demonstrate how lipid-protein interactions can be used to functionally deliver cargo to cells via EVs (III). c Laurdan generalized polarization (GP) of ordered liposomes (LO) composed of 70 mol% DPPC/30 mol% Chol, disordered liposomes (LD) composed of 70 mol% DOPC/30 mol% Chol, HEK293FT cells, giant plasma membrane vesicles (GPMVs) and vesicles from the high-speed centrifugation EV fraction (HS-EVs) and ultracentrifugation EV fraction (UC-EVs) derived from HEK293FT cells. HS-EV and UC-EVs had high Laurdan GP, similar to LO membranes as calculated by Eq. 1. Each dot represents an independent experiment (n ≥ 3). A one-way ANOVA was performed to compare the Laurdan GP of HS-EVs and UC-EVs to the Laurdan GP of all other membranes measured, and comparisons were evaluated using the Sidak multiple comparisons correction. Both EV populations were significantly different from all other vesicle populations except one another (****p < 0.0001). d The frequency at which human membrane-associated proteins and raft proteins are found in EVs as calculated via bioinformatic analysis. Raft associated proteins are more frequently found in EVs compared to a random selection of human membrane-associated proteins. Each dot represents a separate query of 100 human proteins (n = 3). An unpaired, two-tailed t-test was performed to compare the fraction of all proteins and raft associated proteins found in EVs. The fraction of raft associated proteins found in EVs was significantly different from the fraction of all proteins (p < 0.0001). n ≥ 3, error bars represent the standard error of the mean (SEM) throughout the figure. Source data are provided as a Source Data file.

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