Elise Goyet

Shank3 monogenic mutations lead to autism spectrum disorders (ASD). Shank3 is part of the glutamate receptosome that physically links ionotropic NMDA receptors to metabotropic mGlu5 receptors through interactions with scaffolding proteins PSD95–GKAP–Shank3–Homer. A main physiological function of the glutamate receptosome is to control NMDA synaptic function that is required for plasticity induction. Intact glutamate receptosome supports glutamate receptors activation and plasticity induction, while glutamate receptosome disruption blocks receptors activity, preventing the induction of subsequent plasticity. Despite possible impact on metaplasticity and cognitive behaviors, scaffold interaction dynamics and their consequences are poorly defined. Here, we used mGlu5–Homer interaction as a biosensor of glutamate receptosome integrity to report changes in synapse availability for plasticity induction. Combining BRET imaging and electrophysiology, we show that a transient neuronal depolarization inducing NMDA-dependent plasticity disrupts glutamate receptosome in a long-lasting manner at synapses and activates signaling pathways required for the expression of the initiated neuronal plasticity, such as ERK and mTOR pathways. Glutamate receptosome disruption also decreases the NMDA/AMPA ratio, freezing the sensitivity of the synapse to subsequent changes of neuronal activity. These data show the importance of a fine-tuning of protein–protein interactions within glutamate receptosome, driven by changes of neuronal activity, to control plasticity. In a mouse model of ASD, a truncated mutant form of Shank3 prevents the integrity of the glutamate receptosome. These mice display altered plasticity, anxiety-like, and stereotyped behaviors. Interestingly, repairing the integrity of glutamate receptosome and its sensitivity to the neuronal activity rescued synaptic transmission, plasticity, and some behavioral traits of Shank3∆C mice. Altogether, our findings characterize mechanisms by which Shank3 mutations cause ASD and highlight scaffold dynamics as new therapeutic target.

Dopamine receptor D1 modulates glutamatergic transmission in cortico-basal ganglia circuits and represents a major target of L-DOPA therapy in Parkinson’s disease. Here we show that D1 and metabotropic glutamate type 5 (mGlu5) receptors can form previously unknown heteromeric entities with distinctive functional properties. Interacting with Gq proteins, cell-surface D1-mGlu5 heteromers exacerbated PLC signaling and intracellular calcium release in response to either glutamate or dopamine. In rodent models of Parkinson’s disease, D1-mGlu5 nanocomplexes were strongly upregulated in the dopamine-denervated striatum, resulting in a synergistic activation of PLC signaling by D1 and mGlu5 receptor agonists. In turn, D1-mGlu5–dependent PLC signaling was causally linked with excessive activation of extracellular signal–regulated kinases in striatal neurons, leading to dyskinesia in animals treated with L-DOPA or D1 receptor agonists. The discovery of D1-mGlu5 functional heteromers mediating maladaptive molecular and motor responses in the dopamine-denervated striatum may prompt the development of new therapeutic principles for Parkinson’s disease.

Resonance Energy Transfer (RET)-based technologies are used to report protein-protein interactions in living cells. Among them, Bioluminescence-initiated RET (BRET) provides excellent sensitivity but the low light intensity intrinsic to the bioluminescent process hampers its use for the localization of protein complexes at the sub-cellular level. Herein we have characterized the methodological conditions required to reliably perform single-cell BRET imaging using an extremely bright luciferase, Nanoluciferase (Nluc). With this, we achieved an unprecedented performance in the field of protein-protein interaction imaging in terms of temporal and spatial resolution, duration of signal stability, signal sensitivity and dynamic range. As proof-of-principle, an Nluc-containing BRET-based sensor of ERK activity enabled the detection of subtle, transient and localized variations in ERK activity in neuronal dendritic spines, induced by the activation of endogenous synaptic NMDA receptors. This development will improve our comprehension of both the spatio-temporal dynamics of protein-protein interactions and the activation patterns of specific signaling pathways.