An oomycete NLP cytolysin forms transient small pores in lipid membranes.

Microbial plant pathogens secrete a range of effector proteins that damage host plants and consequently constrain global food production. Necrosis and ethylene-inducing peptide 1-like proteins (NLPs) are produced by numerous phytopathogenic microbes that cause important crop diseases. Many NLPs are cytolytic, causing cell death and tissue necrosis by disrupting the plant plasma membrane.

Partitioning of oleic acid into phosphatidylcholine membranes is amplified by strain.

Partitioning of fatty acids into phospholipid membranes is studied on giant unilamellar vesicles (GUVs) utilizing phase-contrast microscopy. With use of a micropipet, an individual GUV is transferred from a vesicle suspension in a mixed glucose/sucrose solution into an isomolar glycerol solution with a small amount of oleic acid added. Oleic acid molecules intercalate into the phospholipid membrane and thus increase the membrane area, while glycerol permeates into the vesicle interior and thus via osmotic inflation causes an increase of the vesicle volume.

The dynamics of melittin-induced membrane permeability.

The transport of co-encapsulated solutes through the melittin-induced pores in the membrane of giant phospholipid vesicles was studied, and the characteristics of the pore formation process were modeled. Molecules of two different sizes (dextran and the smaller, fluorescent marker Alexa Fluor) were encapsulated inside the vesicles. The chosen individual vesicles were then transferred by micromanipulation from the stock suspension to the environment with the melittin (MLT).

A microfluidic diffusion chamber for reversible environmental changes around flaccid lipid vesicles.

The reversible environmental changes around flaccid lipid vesicles represent a considerable experimental challenge, particularly because of remarkable softness of flaccid membranes, which can warp irreversibly under the slightest hydrodynamic flow. As a result, we have developed a microfluidic device for the controlled analysis of individual flaccid, giant lipid vesicles in a changing chemical environment. The setup combines the advantages of a flow-free microfluidic diffusion chamber and optical tweezers, which are used to load the sample vesicles into the chamber.

The response of giant phospholipid vesicles to pore-forming peptide melittin.

The interaction between the pore-forming peptide melittin (MLT) and giant phospholipid vesicles was explored experimentally. Micromanipulation and direct optical observation of a vesicle (loaded with sucrose solution and suspended in isomolar glucose solution) enabled the monitoring of a single vesicle response to MLT. Time dependences of the vesicle size, shape and the composition of the inner solution were examined at each applied concentration of MLT (in the range from 1 to 60 microg/ml).