Frank J.S. Lee

Dual Regulation of NMDA Receptor Functions by Direct Protein-Protein Interactions with the Dopamine D1 Receptor

Dopamine D1-like receptors, composed of D1 and D5 receptors, have been documented to modulate glutamate-mediated fast excitatory synaptic neurotransmission. Here, we report that dopamine D1 receptors modulate NMDA glutamate receptor-mediated functions through direct protein-protein interactions. Two regions in the D1 receptor carboxyl tail can directly and selectively couple to NMDA glutamate receptor subunits NR1-1a and NR2A. While one interaction is involved in the inhibition of NMDA receptor-gated currents, the other is implicated in the attenuation of NMDA receptor-mediated excitotoxicity through a PI-3 kinase-dependent pathway.

Direct binding and functional coupling of {alpha}-synuclein to the dopamine transporters accelerate dopamine-induced apoptosis

Mutations in α-synuclein, a protein highly enriched in presynaptic terminals, have been implicated in the expression of familial forms of Parkinsons disease (PD) whereas native α-synuclein is a major component of intraneuronal inclusion bodies characteristic of PD and other neurodegenerative disorders. Although overexpression of human α-synuclein induces dopaminergic nerve terminal degeneration, the molecular mechanism by which α-synuclein contributes to the degeneration of these pathways remains enigmatic. We report here that α-synuclein complexes with the presynaptic human dopamine transporter (hDAT) in both neurons and cotransfected cells through the direct binding of the non-Aβ amyloid component of α-synuclein to the carboxyl-terminal tail of the hDAT. α-Synuclein-hDAT complex formation facilitates the membrane clustering of the DAT, thereby accelerating cellular dopamine uptake and dopamine-induced cellular apoptosis. Since the selective vulnerability of dopaminergic neurons in PD has been ascribed in part to oxidative stress as a result of the cellular overaccumulation of dopamine or dopamine-like molecules by the presynaptic DAT, these data provide mechanistic insight into the mode by which the activity of these two proteins may give rise to this process.-Lee, F. J. S., Liu, F., Pristupa, Z. B., Niznik, H. B. Direct binding and functional coupling of α-synuclein to the dopamine transporter accelerate dopamine-induced apoptosis.

Dopamine transporter cell surface localization facilitated by a direct interaction with the dopamine D2 receptor.

Altered synaptic dopamine levels have been implicated in several neurological/neuropsychiatric disorders, including drug addiction and schizophrenia. However, it is unclear what precipitates these changes in synaptic dopamine levels. One of the key presynaptic components involved in regulating dopaminergic tone is the dopamine transporter (DAT). Here, we report that the DAT is also regulated by the dopamine D2 receptor through a direct protein–protein interaction involving the DAT amino‐terminus and the third intracellular loop of the D2 receptor. This physical coupling facilitates the recruitment of intracellular DAT to the plasma membrane and leads to enhanced dopamine reuptake. Moreover, mice injected with peptides that disrupt D2–DAT interaction exhibit decreased synaptosomal dopamine uptake and significantly increased locomotor activity, reminiscent of DAT knockout mice. Our data highlight a novel mechanism through which neurotransmitter receptors can functionally modulate neurotransmitter transporters, an interaction that can affect the synaptic neurotransmitter levels in the brain.

FMRP Acts as a Key Messenger for Dopamine Modulation in the Forebrain

The fragile X mental retardation protein (FMRP) is an RNA-binding protein that controls translational efficiency and regulates synaptic plasticity. Here, we report that FMRP is involved in dopamine (DA) modulation of synaptic potentiation. AMPA glutamate receptor subtype 1 (GluR1) surface expression and phosphorylation in response to D1 receptor stimulation were reduced in cultured Fmr1(-/-) prefrontal cortex (PFC) neurons. Furthermore, D1 receptor signaling was impaired, accompanied by D1 receptor hyperphosphorylation at serine sites and subcellular redistribution of G protein-coupled receptor kinase 2 (GRK2) in both PFC and striatum of Fmr1(-/-) mice. FMRP interacted with GRK2, and pharmacological inhibition of GRK2 rescued D1 receptor signaling in Fmr1(-/-) neurons. Finally, D1 receptor agonist partially rescued hyperactivity and enhanced the motor function of Fmr1(-/-) mice. Our study has identified FMRP as a key messenger for DA modulation in the forebrain and may provide insights into the cellular and molecular mechanisms underlying fragile X syndrome.

NCS-1 in the Dentate Gyrus Promotes Exploration, Synaptic Plasticity, and Rapid Acquisition of Spatial Memory

The molecular underpinnings of exploration and its link to learning and memory remain poorly understood. Here we show that inducible, modest overexpression of neuronal calcium sensor 1 (Ncs1) selectively in the adult murine dentate gyrus (DG) promotes a specific form of exploratory behavior. The mice also display a selective facilitation of long-term potentiation (LTP) in the medial perforant path and a selective enhancement in rapid-acquisition spatial memory, phenotypes that are reversed by direct application of a cell-permeant peptide (DNIP) designed to interfere with NCS-1 binding to the dopamine type-2 receptor (D2R). Moreover, the DNIP and the D2R-selective antagonist L-741,626 attenuated exploratory behavior, DG LTP, and spatial memory in control mice. These data demonstrate a role for NCS-1 and D2R in DG plasticity and provide insight for understanding how the DG contributes to the origin of exploration and spatial memory acquisition.

Uncoupling the D1-N-Methyl-D-Aspartate (NMDA) Receptor Complex Promotes NMDA-Dependent Long-Term Potentiation and Working Memory

BACKGROUND Although dopamine D1 receptors are involved in working memory, how D1 receptors contribute to this process remains unclear. Numerous studies have shown that D1 receptors have extensive functional interaction with N-methyl-D-aspartate (NMDA) receptor. Our group previously demonstrated that D1 receptors were able to regulate NMDA receptor functions through direct protein-protein interactions involving the carboxyl terminals of D1 receptors and NMDA receptor NR1a and NR2A subunits respectively. In this study, we explored the effects of the D1-NR1 interaction on NMDA receptor-dependent long-term potentiation (LTP) and working memory by using the TAT-conjugated interfering peptide (TAT-D1-t2). METHODS Miniature excitatory postsynaptic currents are recorded in rat hippocampal primary cultures. Coimmunoprecipitation and calcium/calmodulin-dependent protein kinase II (CaMKII) activity are measured in hippocampal slices and hippocampal neurons under the specified experimental conditions, respectively. Working memory was assessed using a delayed match-to-place protocol in the Morris Water Maze following administration of the TAT-D1-t2 peptide. RESULTS Electrophysiology experiments showed that activation of D1 receptor upregulates NMDA receptor-mediated LTP in a CaMKII-dependent manner. Furthermore, D1 receptor agonist stimulation promotes the NR1-CaMKII coupling and enhances the CaMKII activity; and the D1 receptor-mediated effects can be blocked by the application of the TAT-D1-t2 peptide. Interestingly, animals injected with TAT-D1-t2 peptide exhibited significantly impaired working memory. CONCLUSIONS Our study showed a critical role of NMDA-D1 direct protein-protein interaction in NMDA receptor-mediated LTP and working memory and implicated the involvement of CaMKII in this process.

Direct receptor cross-talk can mediate the modulation of excitatory and inhibitory neurotransmission by dopamine

Dopamine is a neurotransmitter that can regulate both excitatory and inhibitory fast synaptic transmission. The overlapping dopaminergic, glutamatergic, and GABAergic systems provide a basis for the interaction between these three neurotransmitters. Although there is considerable evidence for the involvement of second-messenger systems to mediate receptor cross-talk between these receptor systems, there is emerging evidence that receptors can interact through direct protein-protein interactions. The functional implications and overall significance of the dopamine/glutamate/GABA interactions will be examined.

Genetic factors involved in the pathogenesis of Parkinson's disease.

Parkinsons disease (PD) is a neurodegenerative disease characterized by a loss of nigrostriatal dopaminergic neurons. Recently, PD research has been stimulated by the identification of genes that are implicated in rare familial forms of PD. However, despite these discoveries, the primary cause of PD is still unclear. Various pathogenic mechanisms may be involved including mitochondrial dysfunction, proteasomal dysfunction/protein aggregation, oxidative damage, environmental factors and genetic disposition. Furthermore, dopamine has also been implicated in contributing to the pathogenesis of PD. This review will focus on the genes that have been identified to be associated with PD and how they may impair dopamine metabolism. Understanding the role of these PD-related genes in dopamine neurobiology may provide insight into the underpinning pathogenic mechanisms of PD.

Parkin Disrupts the α-Synuclein/Dopamine Transporter Interaction: Consequences Toward Dopamine-induced Toxicity

Parkinson’s disease is characterized by progressive neuronal degeneration of dopaminergic neurons in the substantia nigra. Many factors are thought to contribute to the neuronal cell death that occurs in Parkinson’s disease, including α-synuclein-mediated toxicity. Previously, we have reported that α-synuclein directly couples to the carboxyl tail of the dopamine transporter (DAT) and that the α-synuclein/DAT protein complex formation accelerates DAT-mediated cellular dopamine (DA) uptake and DA-induced cellular apoptosis. In the present study, we report that parkin, an E2-dependent E3 protein ubiquitin ligase associated with recessive early onset Parkinson’s disease, exerts a protective effect against DA-induced α-synuclein-dependent cell toxicity. Parkin impairs the α-synuclein/DAT coupling by interacting with the carboxyl-terminus of the DAT and blocks the α-synuclein-induced enhancement in both DAT cell surface expression and DAT-mediated DA uptake. Moreover, we have found that parkin protects against DA-induced cell toxicity in dopaminergic SK-N-SH cells. These findings will help identify the role of these proteins in the etiology and/or maintenance of Parkinson’s disease.

Direct interaction between GluR2 and GAPDH regulates AMPAR-mediated excitotoxicity

Over-activation of AMPARs (α−amino-3-hydroxy-5-methylisoxazole-4-propionic acid subtype glutamate receptors) is implicated in excitotoxic neuronal death associated with acute brain insults, such as ischemic stroke. However, the specific molecular mechanism by which AMPARs, especially the calcium-impermeable AMPARs, induce neuronal death remains poorly understood. Here we report the identification of a previously unrecognized molecular pathway involving a direct protein-protein interaction that underlies GluR2-containing AMPAR-mediated excitotoxicity. Agonist stimulation of AMPARs promotes GluR2/GAPDH (glyceraldehyde-3-phosphate dehydrogenase) complex formation and subsequent internalization. Disruption of GluR2/GAPDH interaction by administration of an interfering peptide prevents AMPAR-mediated excitotoxicity and protects against damage induced by oxygen-glucose deprivation (OGD), an in vitro model of brain ischemia.