Stics Analysis Reveals the Role of Tau Phosphorylation in Regulating Lysosome Transport

Abstract

Organelles move through a combination of active transport by motor proteins and diffusion. Tau, a protein that binds to mictrobule tracks, regulates the transport of organelles. To study how tau phosphorylation influences intracellular transport, we applied image correlation spectroscopy analysis to live-cell microscopy images of lysosomes in COS-7 cells expressing tau phosphomutants. Spatio-temporal image correlation spectroscopy (STICS) correlates fluorescence intensity fluctuations within an image through space and over time to measure velocities of mobile molecules and complexes. Compared to conventional tracking analysis, STICS does not rely on detecting and following individual fluorophores through space and time. Using total internal reflection fluorescence microscope, we performed live cell imaging on lysosomes labelled with lysotracker in COS-7 cells, and applied STICS to analyze the image time series. The STICS transport maps were further analyzed to generate histograms of the velocity magnitude and angle distributions of lysosome transport under different tau conditions. Using STICS analysis, we could resolve differences in the direction of lysosome transport under varying tau concentrations. Our results show that lysosome motility towards the cell nucleus increases, in presence of higher tau concentrations, whereas lower concentrations of tau result in vesicles moving both towards the periphery and the perinuclear region. Lysosomes at the central region of the cell are not inhibited by phosphomimetic tau. In contrast, lysosomes at the cell periphery are inhibited by wild-type and phosphomimetic tau. STICS analysis on lysosome motility shows that lysosomes exhibit different motility in the presence of tau according to their localization within the cell. Our results show that tau and its phosphorylation regulate cellular lysosome motility.