Imaging of plant dynamin-related proteins and clathrin around the plasma membrane by variable incidence angle fluorescence microscopy

Endocytosis is an essential phenomenon in eukaryotic cells. In animal cells, dynamin and clathrin play central roles in vesicle formation in the process of endocytosis, but the roles of similar proteins in plants are less well understood. Here, we observed the localization pattern and behavior of GFP-labeled Arabidopsis dynamin-related proteins (DRP1A and DRP2B), and clathrin light chain (AtCLC) around the plasma membrane in tobacco suspension cells by using variable incidence angle fluorescence microscopy (VIAFM). GFP fusions of DRP1A, DRP2B and AtCLC were observed as dot-like puncta 200-500 nm in diameter. The puncta moved to and away from the cell surface or also assembled and disassembled. The localization pattern and behavior of the puncta were similar to those of animal dynamin and clathrin signals reported previously. These results raise the possibility that DRP1A, DRP2B and AtCLC are involved in membrane trafficking around the plasma membrane, including endocytosis.