Abstract
Protein interacting with C kinase (PICK1) is a scaffolding protein that is present in dendritic spines and interacts with a wide array of proteins through its PDZ domain. The best understood function of PICK1 is regulation of trafficking of AMPA receptors at neuronal synapses via its specific interaction with the AMPA GluA2 subunit. Disrupting the PICK1-GluA2 interaction has been shown to alter synaptic plasticity, a molecular mechanism of learning and memory. Lack of potent, selective inhibitors of the PICK1 PDZ domain has hindered efforts at exploring the PICK1-GluA2 interaction as a therapeutic target for neurological diseases. Here, we report the discovery of PICK1 small molecule inhibitors using a structure-based drug design strategy. The inhibitors stabilized surface GluA2, reduced Aβ-induced rise in intracellular calcium concentrations in cultured neurons, and blocked long term depression in brain slices. These findings demonstrate that it is possible to identify potent, selective PICK1-GluA2 inhibitors which may prove useful for treatment of neurodegenerative disorders.
Highlights
Unlike peptides, which have limited cell permeability in the absence of a permeability tag such as a TAT fusion and undesired protein degradation, small molecule inhibitors can be designed for cell-permeability and reduced degradation
We developed a method to assess the importance of pharmacological inhibition of PICK1 on Aβ-mediated changes in synaptic morphology targeting dendritic spine density, using neurons generated from PICK1 KO mice[24]
We developed an assay to determine the functional effect of PICK1 inhibition, by measuring AMPA receptor-dependent intracellular calcium concentrations in neurons cultured from wild-type and PICK1 knockout mice using the FLIPR system as described in Methods
Summary
Its trafficking into and out of the synapse regulates synaptic plasticity and dendritic spine density[9] through interaction of the receptor subunits (GluA1-4) with specific intracellular proteins[10,11,12]. The C-terminus of the GluA2 subunit binds to the PDZ domain of the scaffolding PICK1 protein, an interaction that is required for AMPA receptor internalization and long term depression[13,14,15,16]. A recent study showed that a small molecule inhibitor (BIO922, 1z in this manuscript) of the specific interactions between PICK1 and GluA2 attenuated the effects of Aβ on synapses and surface receptors[20], suggesting that PDZ-domain mediated PICK1 interaction with the GluA2 subunit is required for Aβ effects on synapses and function. The compounds display 200-fold better potency than the endogenous GluA2 peptide ligand, and exhibit unique pharmacological activity in stabilizing neuronal surface GluA2, functionally blocking both Aβ-induced elevation in intracellular calcium concentrations and long term potentiation in cultured neuronal models
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