Activity Dependent Protein Degradation Is Critical for the Formation and Stability of Fear Memory in the Amygdala

Abstract

Protein degradation through the ubiquitin-proteasome system [UPS] plays a critical role in some forms of synaptic plasticity. However, its role in memory formation in the amygdala, a site critical for the formation of fear memories, currently remains unknown. Here we provide the first evidence that protein degradation through the UPS is critically engaged at amygdala synapses during memory formation and retrieval. Fear conditioning results in NMDA-dependent increases in degradation-specific polyubiquitination in the amygdala, targeting proteins involved in translational control and synaptic structure and blocking the degradation of these proteins significantly impairs long-term memory. Furthermore, retrieval of fear memory results in a second wave of NMDA-dependent polyubiquitination that targets proteins involved in translational silencing and synaptic structure and is critical for memory updating following recall. These results indicate that UPS-mediated protein degradation is a major regulator of synaptic plasticity necessary for the formation and stability of long-term memories at amygdala synapses.

Response of the Ubiquitin-Proteasome System to Memory Retrieval After Extended-Access Cocaine or Saline Self-Administration

“…This is analogous to a more established role for the UPS in destabilization of fear memories after their retrieval in a fear conditioning paradigm (Lee et al, 2008;Jarome et al, 2011).…”

“…After this withdrawal period, some rats were given a seeking/retrieval test and then killed 30 or 60 min after memory retrieval (retrieval groups; Figure 2a and b). Timing of the retrieval experiments was based on results from other memory retrieval paradigms (Lee et al, 2008;Jarome et al, 2011;Ren et al, 2013). Other rats were taken from home cages for tissue collection without a seeking test (no retrieval groups) to determine if cocaine withdrawal is associated with altered basal UPS activity and to provide a baseline from which to assess seeking/retrieval test-induced changes in polyubiquitinated protein levels.…”

“…Other rats were taken from home cages for tissue collection without a seeking test (no retrieval groups) to determine if cocaine withdrawal is associated with altered basal UPS activity and to provide a baseline from which to assess seeking/retrieval test-induced changes in polyubiquitinated protein levels. After NAc tissue was harvested, the P2 synaptosomal membrane fraction, which is similar to fractions used in previous work (Lee et al, 2008;Jarome et al, 2011;Ren et al, 2013), was immunoblotted using an antibody recognizing polyubiquitinated proteins targeted for proteasomal degradation (see Materials and Methods). Samples from the retrieval and no retrieval groups were processed for SDS-PAGE and immunoblotting in parallel.…”

“…The UPS-mediated degradation of NSF, a protein whose functions include regulation of AMPA receptor trafficking (Hanley, 2014), has been linked to expression of cocaine CPP (Ren et al, 2013). The synaptic scaffolding protein Shank is tagged and degraded by the UPS after retrieval of fear memories (Lee et al, 2008;Jarome et al, 2011). At the 30-min time point, analysis of NSF levels by two-way ANOVA indicated that NSF was decreased in the retrieval groups compared with no retrieval groups (Figure 2e; main effect: retrieval vs no retrieval: F(1,26) = 5.55, Po0.05).…”

“…This could indicate decreased Shank degradation, although the same rats showed an increase in polyubiquitinated proteins suggestive of a general increase in protein degradation (Figure 2c and d). An alternate possibility is that Shank translation is increased during memory retrieval, as changes in translation and degradation often go hand-in-hand during plasticity (Fonseca et al, 2006;Jarome et al, 2011).…”