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A critical role for PSD-95/AKAP interactions in endocytosis of synaptic AMPA receptors

Abstract

The endocytosis of AMPA receptors (AMPARs) underlies several forms of synaptic plasticity, including NMDA receptor (NMDAR)-dependent long-term depression (LTD), but the molecular mechanisms responsible for this trafficking remain unknown. We found that PSD-95, a major postsynaptic density protein, is important for NMDAR-triggered endocytosis of synaptic AMPARs in rat neuron cultures because of its binding to A kinase–anchoring protein 150 (AKAP150), a scaffold for specific protein kinases and phosphatases. Knockdown of PSD-95 with shRNA blocked NMDAR-triggered, but not constitutive or mGluR-triggered, endocytosis of AMPARs. Deletion of PSD-95's Src homology 3 and guanylate kinase–like domains, as well as a point mutation (L460P), both of which inhibit binding of PSD-95 to AKAP150, also blocked NMDAR-triggered AMPAR endocytosis. Furthermore, expression of a mutant AKAP150 that does not bind calcineurin inhibited this NMDAR-triggered trafficking event. Our results suggest that PSD-95's interaction with AKAP150 is critical for NMDAR-triggered AMPAR endocytosis and LTD, possibly because these scaffolds position calcineurin in the appropriate subsynaptic domain.

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Figure 1: NMDAR- and AMPAR-triggered endocytosis of AMPARs show distinct patterns of internalization.
Figure 2: Acute knockdown of PSD-95 decreases the surface expression and NMDAR-triggered endocytosis of AMPARs.
Figure 3: Constitutive endocytosis and mGluR-triggered endocytosis of AMPARs are not affected by knockdown of PSD-95.
Figure 4: PEST motif deletion and prenylation of PSD-95 does not affect NMDAR-triggered endocytosis of AMPARs.
Figure 5: SH3 and GK domains of PSD-95 are required for NMDAR-triggered endocytosis of AMPARs.
Figure 6: L460P mutation in PSD-95 disrupts binding to AKAP150 and blocks NMDAR-triggered endocytosis of AMPARs.
Figure 7: Binding of calcineurin (PP2B) to AKAP150 is necessary for the NMDAR-triggered endocytosis of AMPARs.
Figure 8: NMDA application causes loss of PSD-95 and AKAP150 from synapses.

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Acknowledgements

We thank A. Ghosh and X. Cai for expert technical assistance and members of the Malenka laboratory for helpful discussions. This work was supported by US National Institutes of Health grant MH63394 (to R.C.M.) and MH080310 (to W.X.) and postdoctoral fellowship SCHL592/4-1 from the DFG (to O.S.).

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All authors contributed to the design and interpretation of the experiments and commented on the manuscript. S.B. conducted and analyzed all experiments. S.B., V.B., W.X. and O.S. prepared constructs. S.B. and R.C.M. wrote the manuscript.

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Correspondence to Robert C Malenka.

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Bhattacharyya, S., Biou, V., Xu, W. et al. A critical role for PSD-95/AKAP interactions in endocytosis of synaptic AMPA receptors. Nat Neurosci 12, 172–181 (2009). https://doi.org/10.1038/nn.2249

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