Neurotransmitters drive combinatorial multistate postsynaptic density networks.

Genes to Cognition, Wellcome Trust Sanger Institute, Cambridgeshire, UK.
Science Signaling (Impact Factor: 7.65). 02/2009; 2(68):ra19. DOI: 10.1126/scisignal.2000102
Source: PubMed

ABSTRACT The mammalian postsynaptic density (PSD) comprises a complex collection of approximately 1100 proteins. Despite extensive knowledge of individual proteins, the overall organization of the PSD is poorly understood. Here, we define maps of molecular circuitry within the PSD based on phosphorylation of postsynaptic proteins. Activation of a single neurotransmitter receptor, the N-methyl-D-aspartate receptor (NMDAR), changed the phosphorylation status of 127 proteins. Stimulation of ionotropic and metabotropic glutamate receptors and dopamine receptors activated overlapping networks with distinct combinatorial phosphorylation signatures. Using peptide array technology, we identified specific phosphorylation motifs and switching mechanisms responsible for the integration of neurotransmitter receptor pathways and their coordination of multiple substrates in these networks. These combinatorial networks confer high information-processing capacity and functional diversity on synapses, and their elucidation may provide new insights into disease mechanisms and new opportunities for drug discovery.

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    ABSTRACT: The C-terminus of AMPA-type glutamate receptor (AMPAR) GluA1 subunits contains several phosphorylation sites that regulate AMPAR activity and trafficking at excitatory synapses. Although many of these sites have been extensively studied, little is known about the signaling mechanisms regulating GluA1 phosphorylation at threonine 840 (Thr840). Here, we report that neuronal depolarization in hippocampal slices induces a calcium and protein phosphatase 1/2A-dependent dephosphorylation of GluA1 at Thr840 and a nearby site at serine 845 (Ser845). Despite these similarities, inhibitors of NMDA-type glutamate receptors and protein phosphatase 2B prevented depolarization-induced Ser845 dephosphorylation but had no effect on Thr840 dephosphorylation. Instead, depolarization-induced Thr840 dephosphorylation was prevented by blocking voltage-gated calcium channels, indicating that distinct Ca2+-sources converge to regulate GluA1 dephosphorylation at Thr840 and Ser845 in separable ways. Results from immunoprecipitation/depletion assays indicate that Thr840 phosphorylation inhibits protein kinase A (PKA)-mediated increases in Ser845 phosphorylation. Consistent with this, PKA-mediated increases in AMPAR currents, which are dependent on Ser845 phosphorylation, were inhibited in HEK-293 cells expressing a Thr840 phosphomimic version of GluA1. Conversely, mimicking Ser845 phosphorylation inhibited protein kinase C phosphorylation of Thr840 in-vitro and PKA activation inhibited Thr840 phosphorylation in hippocampal slices. Together, the regulation of Thr840 and Ser845 phosphorylation by distinct sources of Ca2+ influx and the presence of inhibitory interactions between these sites highlights a novel mechanism for conditional regulation of AMPAR phosphorylation and function.
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