Edited by Michael R. Sussman Keywords:Fluorescent nano bead Clathrin-independent endocytosis Clathrin-dependent endocytosis IKA Wortmannin BY-2 a b s t r a c tTo follow endocytosis in BY-2 cells we made use of fluorescent nano beads. Beads with 20 nm in diameter were internalised rapidly and accumulated partially in compartments also labelled by the endocytic marker FM4-64. Studies in BY-2 cells and protoplasts revealed that larger beads (100 nm) were excluded from uptake into turgescent and plasmolysed cells while protoplasts were able to internalise beads with a diameter of up to 1000 nm. Endocytosis of beads was only partially inhibited by the clathrin-specific inhibitor Ikarugamycin and strongly blocked by wortmannin. These results imply that uptake of beads involves clathrin-dependent and clathrin-independent endocytic mechanisms and supports the hypothesis that clathrin-independent endocytosis plays a general role in plants.

SUMMARYIn eukaryotic cells, several pathways exist for the internalization of plasma membrane proteins and extracellular cargo molecules. These endocytic pathways can be divided into clathrin-dependent and clathrin-independent pathways. While clathrin-dependent pathways are known to be involved in a variety of cellular processes in plants, clathrin-independent pathways have so far only been identified in animal and yeast cells. Here we show that internalization of fluorescent glucose into BY-2 cells leads to accumulation of the sugar in compartments of the endocytic pathway. This endocytic uptake of glucose was not blocked by ikarugamycin, an inhibitor of clathrin-dependent endocytosis, suggesting a role for clathrin-independent endocytosis in glucose uptake. Investigations of fusion and fission of single vesicles by membrane capacitance measurements revealed stimulation of endocytic activity by extracellular glucose. Glucose-stimulated fission of vesicles was not affected by addition of ikarugamycin or blocking of clathrin coat formation by transient over-expression of HUB1 (the C-terminal part of the clathrin heavy chain). These data demonstrate that clathrin-independent endocytosis does occur in plant cells. This pathway may represent a common mechanism for the uptake of external nutrients.

Pressure‐sensitive adhesives based on silicone materials have emerging potential as adhesives in healthcare products, in particular for gentle skin adhesives. To this end, adhesion to rough skin and biocompatibility are crucial factors for a successful implementation. In this study, the mechanical, adhesive, and biological properties of the two‐component poly(dimethylsiloxane) Soft Skin Adhesive MG 7‐9800 (SSA, Dow Corning) have been investigated and compared to Sylgard 184. Different mixing ratios of SSA's components allow for tuning of the shear modulus, thereby modifying the adhesive properties of the polymer. To give a comprehensive insight, the authors have analyzed the interplay between pull‐off stress, adhesion energy, and stretch of the adhesive films on smooth and rough surfaces. The focus is placed on the effects of substrate roughness and on low pressure oxygen plasma treatment of the adhesive films. SSA shows superior biocompatibility in in vitro cell culture experiments. High pull‐off stresses in the range of 3 N cm−2 on a rough surface are achieved, promising broad application spectra for SSA‐based healthcare products.

To analyze the kinetics and size of single exo- and endocytotic events in BY-2 protoplasts, we employed cell-attached membrane capacitance measurements. These measurements revealed different modes of fusion and fission of single vesicles. In about half of the observed exocytotic events, fusion occurred transiently, which facilitates rapid recycling of vesicles. In addition, transient sequential or multi-vesicular exocytosis observed in some recordings can contribute to an increase in efficiency of secretory product release. Microscopic analysis of the timescale of cellulose and pectin deposition in protoplasts demonstrates that rebuilding of the cell wall starts soon after isolation of protoplasts and that transient fusion events can fully account for secretion of the required soluble material. The capacitance measurements also allowed us to investigate formation of the fusion pore. We speculate that regulation of secretion may involve control of the length and/or size of fusion pore opening. Together, the different kinetic modes of exo- and endocytosis revealed by capacitance measurements underline the complexity of this process in plants and provide a basis for future research into the underlying mechanisms. The fact that similar fusion/fission kinetics are present in plant and animal cells suggests that many of these mechanisms are highly conserved among eukaryotes.

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