UC San Diego

Changes in synaptic strength underlie the basis of learning and memory and are controlled, in part, by the insertion and removal of AMPA-type glutamate receptors. Once internalized, these receptors maybe recycled back to the plasma membrane by subunit-specific interactions with other proteins or by post-translational modifications such as phosphorylation. Ubiquitination, another post-translational modification, has recently emerged as a key signal that regulates the recycling and trafficking of glutamate receptors. However, the activity-dependent rules that govern internalization and degradation have not been fully elucidated.

This dissertation was designed in two parts; (1) to investigate the molecular specificity and activity-dependent rules of AMPAR ubiquitination via a deubiquinating enzyme, USP8 and (2) to determine how the lysosome, an organelle that degrades AMPARs, may also respond to synaptic cues to coordinate degradation of membrane proteins such as AMAPRs. We approached the first part of this study by evaluating the distribution and activation of USP8 and how neuronal activity alters USP8 distribution and activation. Using mainly biochemical techniques such as western blot and immunopreciptation as well as immunostaining, we showed that NMDAR signaling is required to spare AMPARs from ubiquitin-dependent internalization and degradation. To elucidate activity-dependent trafficking of lysosomes, we utilized electron microscopy and confocal live imaging of neurons and 2-photon imaging of organotypic hippocampal slices to visualize lysosomes. We found that lysosomes are positioned locally at dendritic spines and can be recruited do a dendritic spine in an activity-dependent manner.

These findings provide evidence for activity-dependent control of a DUB, USP8, and an organelle, the lysosome, at synapses. We showed the dynamic regulation of USP8 is critical in maintaining synapses and scaling during homeostatic plasticity and that neuronal activity can signal to the lysosome to locally degrade membrane proteins. These studies suggest that localized degradation can play an active role in remodeling synapses to facilitate plastic events and help locally maintain cellular homeostasis.