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 Aß-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.
Subject(s)
Amyloid beta-Peptides/metabolism , Brain/metabolism , Carrier Proteins/antagonists & inhibitors , Dendritic Spines/metabolism , Neurodegenerative Diseases/metabolism , Nuclear Proteins/antagonists & inhibitors , Synapses/metabolism , Animals , Brain/pathology , Calcium/metabolism , Calcium Signaling , Carrier Proteins/metabolism , Cell Cycle Proteins , Dendritic Spines/pathology , Drug Design , Mice , Neurodegenerative Diseases/drug therapy , Neurodegenerative Diseases/pathology , Nuclear Proteins/metabolism , PDZ Domains , Receptors, AMPA/metabolism , Synapses/pathologyABSTRACT
The membrane protein interacting with kinase C1 (PICK1) plays a trafficking role in the internalization of neuron receptors such as the amino-3-hydroxyl-5-methyl-4-isoxazole-propionate (AMPA) receptor. Reduction of surface AMPA type receptors on neurons reduces synaptic communication leading to cognitive impairment in progressive neurodegenerative diseases such as Alzheimer disease. The internalization of AMPA receptors is mediated by the PDZ domain of PICK1 which binds to the GluA2 subunit of AMPA receptors and targets the receptor for internalization through endocytosis, reducing synaptic communication. We planned to block the PICK1-GluA2 protein-protein interaction with a small molecule inhibitor to stabilize surface AMPA receptors as a therapeutic possibility for neurodegenerative diseases. Using a fluorescence polarization assay, we identified compound BIO124 as a modest inhibitor of the PICK1-GluA2 interaction. We further tried to improve the binding affinity of BIO124 using structure-aided drug design but were unsuccessful in producing a co-crystal structure using previously reported crystallography methods for PICK1. Here, we present a novel method through which we generated a co-crystal structure of the PDZ domain of PICK1 bound to BIO124.
Subject(s)
Carrier Proteins/chemistry , Carrier Proteins/metabolism , Nuclear Proteins/chemistry , Nuclear Proteins/metabolism , Small Molecule Libraries/chemistry , Small Molecule Libraries/pharmacology , Binding Sites/drug effects , Crystallography , Drug Design , Humans , Models, Molecular , Molecular Conformation , PDZ Domains , Protein Binding/drug effects , Receptors, AMPA/metabolism , Structure-Activity RelationshipABSTRACT
Amyloid beta (Aß), a key component in the pathophysiology of Alzheimer's disease, is thought to target excitatory synapses early in the disease. However, the mechanism by which Aß weakens synapses is not well understood. Here we showed that the PDZ domain protein, protein interacting with C kinase 1 (PICK1), was required for Aß to weaken synapses. In mice lacking PICK1, elevations of Aß failed to depress synaptic transmission in cultured brain slices. In dissociated cultured neurons, Aß failed to reduce surface α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor subunit 2, a subunit of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors that binds with PICK1 through a PDZ ligand-domain interaction. Lastly, a novel small molecule (BIO922) discovered through structure-based drug design that targets the specific interactions between GluA2 and PICK1 blocked the effects of Aß on synapses and surface receptors. We concluded that GluA2-PICK1 interactions are a key component of the effects of Aß on synapses.
Subject(s)
Amyloid beta-Peptides/toxicity , Carrier Proteins/metabolism , Excitatory Postsynaptic Potentials , Nuclear Proteins/metabolism , Peptide Fragments/toxicity , Synapses/metabolism , Animals , Carrier Proteins/genetics , Cell Cycle Proteins , Cells, Cultured , Hippocampus/cytology , Hippocampus/metabolism , Hippocampus/physiology , Mice , Neurons/drug effects , Neurons/metabolism , Neurons/physiology , Nuclear Proteins/genetics , Protein Binding , Rats , Receptors, AMPA/metabolism , Synapses/drug effects , Synapses/physiologyABSTRACT
Tendons are collagen-based fibrous tissues that connect and transmit forces from muscle to bone. These tissues, which are high in collagen type I content, have been studied extensively to understand collagen fibrillogenesis. Although the mechanisms have not been fully elucidated, our understanding has continued to progress. Here, we review two prevailing models of collagen fibrillogenesis and discuss the regulation of the process by candidate cellular and extracellular matrix molecules. Although numerous molecules have been implicated in the regulation of collagen fibrillogenesis, we focus on those that have been suggested to be particularly relevant to collagen type I fibril formation during tendon development, including members of the collagen and small leucine-rich proteoglycan families, as well as other molecules, including scleraxis, cartilage oligomeric matrix protein, and cytoskeletal proteins.
