Axonal transport of the ubiquitin-proteasome system and it relevance for protein degradation

OTERO MG; CROMBERG L; FALZONE T

Paris

Conferencia; International Conference on Alzheimer Disease; 2011

Alzhimer Association

Aberrant accumulation of misfolded protein in AD suggests that protein degradation defects by the Ubiquitin Proteasome System (UPS) are implicated in disease pathogenesis. Amyloid dependent impaired UPS degradation was observed in brain areas suggesting that UPS localization is crucial for proper protein removal. Consequently, it is important to understand how UPS is moved along axons and how defects in UPS delivery can lead to dysfunction in protein degradation as is observed in AD. Methods To understand the axonal transport properties of proteasome subunits we generated an expression vector driving PSMA-7 proteasome subunit fused to YFP. To unravel if neuronal proteasomes can be transported associated with membrane compartment we performed flotation experiments in continuous sucrose density gradients. To understand if proteasome function can regulate its own axonal transport we perform PSMA-YFP movies in the presence of proteasome inhibition. Results In vivo imaging of transfected primary neurons using PSMA7-YFP revealed high amount of particulate erratic PSMA7-YFP movement and a proportion of both anterograde (10,11%) and retrograde (12,68%) processive movement. Interestingly, anterograde PSMA7-YFP display average speeds of 0,187 um/sec and a retrograde speed of 0,771 um/sec. Bottom loaded flotation experiments suggested that UPS is enriched in soluble fractions. However, UPS floats between fractions 5-13 suggesting an association to a membrane compartment. Blocking the UPS by MG132 induced significant changes in PSMA7-YFP transport proportions. Significant and selective decrease in retrograde proportion with an increase in stationary particles was observed and changes in both anterograde and retrograde speed were observed after MG132. Conclusions The UPS is actively transported in axons showing particulate processive movement in both anterograde and retrograde direction. Sucrose density flotation experiments indicate that a fraction of neuronal proteasome can be associated with membranes. Interestingly, inhibition of the proteasome induced selective reductions in retrograde UPS transport suggesting that proteasome activity can modulate its own transport dynamics. Taken together, these experiments suggest a relevant function for the axonal transport of the UPS and we can propose that defects in the transport system can lead to abnormal protein accumulation similar to what is observed in neurodegenerative diseases such as Alzheimer Disease.