Novel axonal transport motion regimes of the proteasome complex analyzed by newly desigened tracking system

M ALLOATTI; MG OTERO; LE CROMBERG; VM POZO DEVOTO; TMM SAEZ; L BRUNO; TL FALZONE

Huerta Grande

Congreso; XXVIII Congreso de la Sociedad Argentina de Neurociencias; 2013

SAN

While the ubiquitin-proteasome system (UPS) is the major route for cytosolic protein degradation, local accumulations of poly-ubiquitinated proteins in neurodegenerative diseases suggest that protein turn over by the proteasome might be impaired during Alzheimer disease (AD). Local clearance by the UPS depends on the correct delivery and positioning of the proteasome complex. Our previous results showed UPS transport throughout axons; however, proteasome distribution was not fully understood. We performed live imaging experiments of fluorescent proteasomes in primary hippocampal neurons. High temporal resolution movies (50 frames per second) were analyzed with a high spatial resolution tracking system allowing the subpixel identification of particle position. Through a set of computer scripts we tracked individual proteasome particles and based on the mean square displacement we classified them in three motion categories: actively transported diffusive and confined motion. We revealed from active proteasomes a distribution of segmental velocities according to molecular motor-dependent transport. Proteasome diffusion coefficient was obtained and was consistent with proteasome size. Finally a cage of confinement was obtained for restricted movement. This new approach allowed the characterization of at least three proteasome motion regimes that support proteasome distribution in axons and might provide clues about the defects associated with local protein accumulation in AD.