A binding site outside the canonical PDZ domain determines the specific interaction between Shank and SAPAP and their function

Abstract

Shank and SAPAP (synapse-associated protein 90/postsynaptic density-95-associated protein) are two highly abundant scaffold proteins that directly interact with each other to regulate excitatory synapse development and plasticity. Mutations of SAPAP, but not other reported Shank PDZ domain binders, share a significant overlap on behavioral abnormalities with the mutations of Shank both in patients and in animal models. The molecular mechanism governing the exquisite specificity of the Shank/SAPAP interaction is not clear, however. Here we report that a sequence preceding the canonical PDZ domain of Shank, together with the elongated PDZ BC loop, form another binding site for a sequence upstream of the SAPAP PDZ-binding motif, leading to a several hundred-fold increase in the affinity of the Shank/SAPAP interaction. We provide evidence that the specific interaction afforded by this newly identified site is required for Shank synaptic targeting and the Shank-induced synaptic activity increase. Our study provides a molecular explanation of how Shank and SAPAP dosage changes due to their gene copy number variations can contribute to different psychiatric disorders.Shank | SAPAP | PDZ | synapse | specific interaction P ostsynaptic density (PSD) at excitatory synapses refers to disk-shaped, densely packed mega-protein assemblies located beneath postsynaptic membranes (1-5). A set of highly abundant scaffold proteins, including DLGs (disc large proteins including PSD-95, PSD-93, and SAP102), SAPAP (synapse-associated protein 90/postsynaptic density-95-associated protein; also known as GKAP or DLGAP), and Shank, are known to be critical for the formation, stability, and neuronal activity-dependent dynamic regulations of . In addition to orchestrating PSD formation, these scaffold proteins also serve to control the trafficking and clustering of receptors on the plasma membranes and to interface with actin cytoskeletons at synapses (15,18,19). Mutations of DLGs, SAPAP, and Shank have been found to cause or associate with various psychiatric disorders, including autism spectrum disorder (ASD), depression, and schizophrenia (20-28), further supporting the importance of these proteins in normal brain development and functions.Electron and superresolution light microscopy imaging studies (4,5,18,(29)(30)(31)(32) have revealed that proteins in PSDs form distinct layers along the axodendritic axis of synapses with a sequential order of membrane-spanning glutamate receptors and cell adhesion molecules, DLGs, the SAPAP and Shank scaffolds, and actin cytoskeletons. Such a distinct layered structure in PSDs depends critically on the specific interactions between the scaffold proteins. For example, the phosphorylation-dependent interaction between DLG GK domain and the N-terminal repeating sequences of SAPAP positions SAPAP beneath DLGs (33), and the interaction between the SAPAP PDZ-binding motif (PBM) and Shank PDZ domain functions to place Shank at a deeper layer in PSD (7).The critical roles of the PSD scaffold proteins in norma...

Structural basis for PDZ domain interactions in the post‐synaptic density scaffolding protein Shank3

“…In line with this, secondary structure predictions suggest one short helix and two β strands in the linker – the latter were also seen in the complex between Shank3 and an extended GKAP peptide (Zeng et al . ).…”

“…), as well as for an extended GKAP peptide in complex with the PDZ domain of Shank3 (Zeng et al . ).…”

Phase Transition in Postsynaptic Densities Underlies Formation of Synaptic Complexes and Synaptic Plasticity

“…Isothermal titration calorimetry (ITC)-based titration assay showed that N-terminal thioredoxin-tagged last 30 residues of SynGAP (SynGAP PBM) binds to each of the three PDZ domains with comparable and weak affinities (K d of a few dozens of micromolars; Figure 1G, G1). Such weak bindings are unlikely to support a specific functional interaction between SynGAP and PSD-95 in vivo (Ye and Zhang, 2013; Zeng et al, 2016). …”

PDZ Domains as Drug Targets

“…The variant Arg536 (R536W) is positioned in the linker region between the SH3 domain and the PDZ domain of Shank3, but does not interfere with the binding of C-terminal ligands nor changes the overall structure of the SH3-PDZ tandem. [40] The implication of Shank3 in neurological disorder has promoted the development of inhibitors toward its PDZ domain. The PDZ domains of the Shank proteins are members of the class I PDZ domains, with a His positioned in the αB1 helix, which is proposed to bind primarily C-terminal ligands with a S/T-x-ϕ motif (Figure 8b,c).…”

“…[2] Several www.advancedsciencenews.com www.advtherap.com Figure 2. [40] In yet further distinctive binding mode variations of the canonical interaction, P 0 is buried deeper in the binding groove of the PDZ domain, occupying what corresponds to the P 1 position (Figure 2e). In a canonical insertion, there is a hydrogen bonding network (orange dashed lines) between the backbone of the PDZ domain and the backbone of the peptide.…”