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Anna Hupalowska (International Institute of Molecular and Cell Biology, Warsaw, Poland)
The mechanism of endocytosis comprises selection of cargo at the plasma membrane, formation of vesicles and their move towards a series of intracellular compartments. Having reached early endosomes, internalized ligand-receptor complexes can be recycled to the cell surface via recycling endosomes or directed to degradative compartments like late endosomes and lysosomes. In our studies we focused on the early stages of endocytosis, in particular on APPL-positive endosomes. This subpopulation of early endosomes appears physically separated from the canonical early endosomes harboring Rab5 and its other effector EEA1. In order to discover the properties and the function of APPL endocytic compartment we established the kinetics of cargo transit through APPL endosomes in comparison to EEA1 endosomes using microscopical approaches. After taking confocal images of APPL1, EEA1 and internalized cargo we performed a global and quantitative analysis of endosomes using vesicle tracking algorithms (MotionTracking software) developed by Dr. Yannis Kalaidzidis. Following automated object recognition, further statistical analyses of vesicles (size, number, fluorescence intensity, colocalization wi