Effect of G4C2 repeat expansions on the motion of lysosomes inside neurites

The G4C2hexanucleotide repeat expansion (HRE) in the c9orf72 locus is a mutation associated with amyotrophic lateral sclerosis. Recent evidence suggests a link with disrupted axonal trafficking in neurons. Here, using a neuronal-like cell line without or transfected with G4C2repeats, we characterize the motion of lysosomes inside neurites. The neurites grew either aligned to patterned lines, or oriented freely on a 2D-substrate. Implementing time-resolved (local) mean squared displacement analysis lysosome trajectories were split into sub-diffusive, diffusive, and super-diffusive parts. Our results suggest that in the presence of the G4C2repeats, lysosome trafficking is hampered, exhibiting overall decreased mean squared displacement and speed, more prominently inside aligned neurites. Moreover, a prominent effect in the super-diffusive drift velocity and diffusive motion diffusion coefficient was evident when the motion occurred inside aligned neurites. Trajectories which included super-diffusive motion, exhibited a varied ratio of anterograde/retrograde/neutral for both neurite geometries in the presence of G4C2repeats but a similar velocity decrease for both directions in each neurite geometry. Our findings support the hypothesis that impaired axonal trafficking emerges in the presence of the G4C2HRE, and demonstrate that this effect is more prominent when the neurites are aligned.