Abstract
One of the core pathogenic mechanisms for schizophrenia is believed to be dysfunction in glutamatergic synaptic transmissions, particularly hypofunction of N-methyl d-aspartate receptors (NMDARs). Previously we showed that 14-3-3 functional knockout mice exhibit schizophrenia-associated behaviors accompanied by reduced synaptic NMDARs in forebrain excitatory neurons. To investigate how 14-3-3 proteins regulate synaptic localization of NMDARs, here we examined changes in levels of synaptic NMDARs upon 14-3-3 inhibition in primary neurons. Expression of 14-3-3 protein inhibitor (difopein) in primary glutamatergic cortical and hippocampal neurons resulted in lower number of synaptic puncta containing NMDARs, including the GluN1, GluN2A, or GluN2B subunits. In heterologous cells, 14-3-3 proteins enhanced surface expression of these NMDAR subunits. Furthermore, we identified that 14-3-3ζ and ε isoforms interact with NMDARs via binding to GluN2A and GluN2B subunits. Taken together, our results demonstrate that 14-3-3 proteins play a critical role in NMDAR synaptic trafficking by promoting surface delivery of NMDAR subunits GluN1, GluN2A, and GluN2B. As NMDAR hypofunctionality is known to act as a convergence point for progression of symptoms of schizophrenia, further studies on these signaling pathways may help understand how dysfunction of 14-3-3 proteins can cause NMDAR hypofunctionality and lead to schizophrenia-associated behaviors.
Highlights
N-methyl-d-aspartate receptors (NMDARs) are ionotropic glutamate receptors that play a central role in excitatory synaptic transmission and plasticity [1, 2]
We demonstrate that inhibition of 14-3-3 proteins leads to decreased synaptic localization of NMDAR subunits GluN1, GluN2A, and GluN2B in primary cultures of glutamatergic cortical and hippocampal neurons
Results from both surface biotinylation and ICC show that 14-3-3 proteins enhance surface expression of these NMDAR subunits in heterologous cells
Summary
N-methyl-d-aspartate receptors (NMDARs) are ionotropic glutamate receptors that play a central role in excitatory synaptic transmission and plasticity [1, 2]. They form heteromeric assemblies composed of GluN1, GluN2, and GluN3 subunits, which contain a large amino (N)-terminal extracellular domain, three membrane-spanning domains, a ‘hairpin’ loop that forms the pore-lining region, and an intracellular carboxy (C)-terminal domain [2]. NMDARs have a voltage-sensitive Mg2+ block and high permeability to Ca2+. This Ca2+ influx triggers signal transduction cascades that regulate synaptogenesis, experience-dependent synaptic remodeling, and long-lasting changes in synaptic efficacy such as long-term potentiation and long-term depression [1, 3, 4].
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