Abstract
The protein machinery controlling membrane fusion (or fission) has been well studied; however, the role of vesicle diffusion near membranes in these critical processes remains unclear. We experimentally and theoretically investigated the dynamics of small vesicles (∼50nm in diameter) that are diffusing near supported planar bilayers acting as “target” membranes. Using total internal reflection-fluorescence correlation spectroscopy, we examined the validity of theoretical analyses of vesicle–membrane interactions. Vesicles were hindered by hydrodynamic drag as a function of their proximity to the planar bilayer. The population distributions and diffusion kinetics of the vesicles were further affected by changing the ionic strength and pH of the buffer, as well as the lipid composition of the planar membrane. Effective surface charges on neutral bilayers were also analyzed by comparing experimental and theoretical data, and we show the possibility that vesicle dynamics can be modified by surface charge redistribution of the planar bilayer. Based on these results, we hypothesize that the dynamics of small vesicles, diffusing close to biomembranes, may be spatially restricted by altering local physiological conditions (e.g., salt concentration, lipid composition, and pH), which may represent an additional mechanism for controlling fusion (or fission) dynamics.
Highlights
Small vesicles (,100 nm in diameter) found inside cells participate in vital cellular processes such as exocytosis and endocytosis [1,2,3,4,5]
Vesicles typically undergo two stages of dynamics, and for simplicity, we focus on the intermembrane interactions occurring before exocytosis: vesicles first must diffuse in close proximity with the target membrane; the vesicles dock and fuse with membranes via specific protein-protein interactions to release their contents [6]. (The steps for the intermembrane interactions occurring during postendocytotic fission are reversed.) The interactions of vesicles with membranes and their regulation at the second stage have been investigated widely using model systems and living cells [7,8]
Increasing ionic strength slows vesicle dynamics near target supported planar membranes To evaluate the effect of ionic strength on intermembrane interactions, we used negatively charged small unilamellar vesicles (SUVs) and planar membranes that mimic a plasma membrane target
Summary
Small vesicles (,100 nm in diameter) found inside cells participate in vital cellular processes such as exocytosis and endocytosis [1,2,3,4,5] During these processes, vesicles typically undergo two stages of dynamics, and for simplicity, we focus on the intermembrane interactions occurring before exocytosis: vesicles first must diffuse in close proximity with the target membrane; the vesicles dock and fuse with membranes via specific protein-protein interactions to release their contents [6]. Because the cytoplasmic leaflet of the plasma membrane is enriched in anionic lipid (;10–20 mol %) [9], electrostatic repulsion between opposing membranes is a primary intermembrane interaction, as well as interactions mediated by cytosolic calcium. Some work has discussed hydrodynamic interactions of nanoparticles with a surface, but these measurements suffered from low signal/noise ratio when the particle diameter was reduced to ;50 nm [23,24]
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