Differential Regulation of AMPA Receptor Trafficking by Neurabin-Targeted Synaptic Protein Phosphatase-1 in Synaptic Transmission and Long-Term Depression in Hippocampus

Abstract

Filamentous actin binding protein neurabin I (NrbI) targets protein phosphatase-1 (PP1) to specific postsynaptic microdomains, exerting critical control over AMPA receptor (AMPAR)-mediated synaptic transmission. NrbI-targeted synaptic PP1, which promotes synaptic depression upon long-term depression (LTD) stimuli, serves to prevent synaptic depression under basal conditions. The present studies investigate this opposite regulation of AMPAR trafficking during basal synaptic transmission and LTD by expressing NrbI or NrbI mutant, which is defective in PP1 binding, in hippocampal slice or neuron cultures. We find that expression of the NrbI mutant to interfere with PP1 targeting dramatically reduces basal synaptic transmission, which is correlated with the reduction in surface expression of AMPA subtype glutamate receptor (GluR) 1 and GluR2 subunits. Biochemical analysis demonstrates that the NrbI mutant selectively increases the phosphorylation of GluR2 at C-terminal consensus PKC site, serine 880, which is known to favor GluR2 interaction with PDZ (postsynaptic density 95/Discs large/zona occludens 1) protein PICK1 (protein interacting with C kinase-1). Inhibition of PKC activity or GluR2-PICK1 interaction completely reverses the synaptic depression in neurons expressing the NrbI mutant, suggesting that NrbI-targeted synaptic PP1 stabilizes the basal transmission by negatively controlling PKC phosphorylation of GluR2 and the subsequent PICK1-mediated decrease in GluR2-containing AMPAR surface expression. Distinct from basal transmission, blocking GluR2-PICK1 interaction or PKC activity produces minimal effects on LTD in NrbI-expressing neurons. Instead, NrbI-targeted PP1 facilitates LTD by dephosphorylating GluR1 at both serine 845 and serine 831, with GluR2 serine 880 phosphorylation unaltered. Our studies thus elucidate that NrbI-targeted PP1, in response to distinct synaptic activities, regulates the synaptic trafficking of specific AMPAR subunits.

Rab8 Interacts with Distinct Motifs in α2B- and β2-Adrenergic Receptors and Differentially Modulates Their Transport

“…After 10 days in vitro, GFP-Rab8 constructs were transfected into the neurons with the calcium phosphate precipitation method as described previously (42). Two days after transfection, the neurons were stained with antibodies recognizing ␣ 2B -AR at a dilution of 1:50 for 1 h at room temperature followed by incubation with Alexa 568-conjugated secondary antibodies for 1 h. Images were acquired using a confocal microscope (Zeiss 510 Meta) as described (43). Data were analyzed by NIH Image J software.…”

Synaptic NMDA receptor stimulation activates PP1 by inhibiting its phosphorylation by Cdk5

“…For example, the serine/threonine protein phosphatase PP1 is enriched in dendritic spines at excitatory synapses (Ouimet et al, 1995;Strack et al, 1999;Terry-Lorenzo et al, 2000), and it controls synaptic plasticity through its ability to dephosphorylate important substrates at the synapse, including Ser845 of the GluA1 -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor, and Thr286 of CaM kinase II (Bito et al, 1996;Strack et al, 1997;Genoux et al, 2002;Hsieh-Wilson et al, 2003;Hu et al, 2007), and substrates in the nucleus such as the transcription factor CREB (Bito et al, 1996).…”

Spinophilin-Targeted Protein Phosphatase-1 Alleviated Inflammatory Pain by Negative Control of MEK/ERK Signaling in Spinal Cord Dorsal Horn of Rats

“…However, the synaptic strength is definitely altered when PP1 interaction with some of these scaffolding proteins is selectively disrupted (Yan et al, 1999;Hu et al, 2007), suggesting that PP1, targeted by different scaffolding proteins, might play distinct or even opposite roles in synaptic responses. In acutely dissociated striatal or neostriatal neurons, disturbing SPN/PP1 interaction blocks the rundown of AMPAR-mediated inward currents generated by application of exogenous kainate (Yan et al, 1999;Feng et al, 2000).…”

“…The supernatant (S1) was collected and centrifuged at 10,000 ϫ g for 15 min to obtain P2 pellet that contained crude synaptosomal fraction. P2 was incubated for 30 min with the lysis buffer containing 0.5% Triton X-100, and then centrifuged at 32,000 ϫ g for 20 min to harvest synaptosomal membrane fraction (P3; Smith et al, 2006;Jaworski et al, 2009;Sanz-Clemente et al, 2010;Li et al, 2015). To assay the protein expression and phosphorylation (Li et al, 2015), the spinal dorsal horn was homogenized in radioimmunoprecipitation assay (RIPA) buffer (50.0 mM Tris-HCl, pH 8.0, 150.0 mM NaCl, 1.0 mM EDTA, 1.0% Nonidet P-40, 0.1% SDS, 0.5% sodium deoxycholate, and proteases/phosphatases inhibitors).…”

“…The paired-pulse ratios were measured by delivering two stimuli at different interstimulus intervals (ISIs; Jurado et al, 2013). The series and input resistances were monitored on-line throughout each experi- (Hu et al, 2007). The recordings were abandoned if any resistance changed Ͼ15%.…”