Biomimetic divalent ligands for the acute disruption of synaptic AMPAR stabilization

Abstract

The interactions of the AMPA receptor (AMPAR) auxiliary subunit Stargazin with PDZ domain-containing scaffold proteins such as PSD-95 are critical for the synaptic stabilization of AMPARs. To investigate these interactions, we have developed biomimetic competing ligands that are assembled from two Stargazin-derived PSD-95/DLG/ZO-1 (PDZ) domain-binding motifs using 'click' chemistry. Characterization of the ligands in vitro and in a cellular FRET-based model revealed an enhanced affinity for the multiple PDZ domains of PSD-95 compared to monovalent peptides. In cultured neurons, the divalent ligands competed with transmembrane AMPAR regulatory protein (TARP) for the intracellular membrane-associated guanylate kinase resulting in increased lateral diffusion and endocytosis of surface AMPARs, while showing strong inhibition of synaptic AMPAR currents. This provides evidence for a model in which the TARP-containing AMPARs are stabilized at the synapse by engaging in multivalent interactions. In light of the prevalence of PDZ domain clusters, these new biomimetic chemical tools could find broad application for acutely perturbing multivalent complexes.

Reduced PDZ Interactions of Rescued ΔF508CFTR Increases Its Cell Surface Mobility

“…For instance, stimulated G-protein coupled receptors that contain PDZ-binding domains segregate into specific clathrin-coated pits to greatly extend their surface lifetime through actin association (39). Similar to the present study, a correlative analysis of protein diffusion and internalization has been performed for the neuronal ␣-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) (40). The AMPAR forms a complex with Stargazin and the PDZ scaffold PSD-95, which result in receptor immobilization at the neuronal postsynaptic density.…”

“…The AMPAR forms a complex with Stargazin and the PDZ scaffold PSD-95, which result in receptor immobilization at the neuronal postsynaptic density. Disruption of Stargazin-PSD-95 interactions with a cell permeable peptide mimetic of the Stargazin PDZ binding domain increased AMPAR diffusive range by ϳ1.3-fold and endocytosis by ϳ4-fold (40). For CFTR, Thelin et al (41) reported that the S13F mutation increased CFTR diffusion coefficient by only ϳ20%, but increased internalization by ϳ2-fold.…”

Molecular basis of the alternative recruitment of GABAA versus glycine receptors through gephyrin

“…This principle has been demonstrated for the interaction between the N-methyl-D-aspartate-type glutamate receptors and the scaffolding postsynaptic density protein 95 (PSD-95), which has been targeted by peptide-based inhibitors [47][48][49][50][51][52] . This approach has shown great promise both as a pharmacological tool 51,53 and, in particular, in the development of therapeutically relevant compounds 54,55 . We therefore propose that conceptually similar molecules could be used to interfere with the receptor-scaffold interactions in vivo to modulate GABAergic and/or glycinergic transmission.…”

Fluorescent Amino Acids: Modular Building Blocks for the Assembly of New Tools for Chemical Biology

“…38 These phthalimide and naphthalimide-based FlAAs have also been incorporated into peptides recognizing PDZ and SH2 domains revealing their versatility in diverse applications. [39][40][41][42] The success of the dimethylaminonaphthalimide family led to their further exploration as environment-sensitive FlAAs. For example, the 4-(N,Ndimethylamino)naphthalimide amino acid (4-DMNA) was developed to complement 4-DAPA and 6-DMNA, and also yielded excellent improvements in stability towards hydrolysis and ease of synthesis.…”