Cytoskeletal protein tau dynamics as determined by photoactivation and pixel analysis

CARINA WEISSMANN; HANS JÜRGEN REYHER; HEINZ JÜRGEN STEINHOFF; ROLAND BRANDT

Viena, Austria

Congreso; Fens 5th Forum of European Neuroscience; 2006

Tau is a cytoskeletal protein that is mainly enriched in axons where it is thought to regulate microtubule dynamics. Tau is also involved in many diseases referred to as "tauopathies" including Alzheimer´s disease where tau forms neurofibrillary tangles via unknown mechanisms. In order to investigate tau dynamics in living cells, fusion constructs between human tau and photoactivatable green fluorescent protein (PA-GFP; Patterson and Lippincott-Schwartz, 2002) were prepared. The constructs were transfected in neural PC12 cells to obtain clonal lines stably expressing the fusion proteins. A semi-automatic system was set up using a laser scanning confocal microscope (LSCM) equipped with an incubation chamber and 488nm and 405 nm lasers. To measure the mobility of these fusion proteins within living cells time lapse analysis was performed and individual images collected. The image series were further processed with a pixel analysis program specifically developed for this purpose. Preliminary data analysis suggests that the mobility of tau follows a free diffusion model. Diffusion coefficients determined with this method may allow to analyze different interactions occurring between proteins within living cells and how these interactions are affected under physiological conditions. Photoactivation and time lapse analysis of cytoskeletal proteins provides a useful tool to analyze cytoskeletal dynamics in living cells. The approach permits to calculate diffusion coefficients of cellular proteins.