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Adapting the endoplasmic reticulum proteostasis rescues epilepsy-associated NMDA receptor variants

Acta Pharmacologica Sinica volume 45, pages 282–297 (2024)Cite this article

Abstract

The GRIN genes encoding N-methyl-D-aspartate receptor (NMDAR) subunits are remarkably intolerant to variation. Many pathogenic NMDAR variants result in their protein misfolding, inefficient assembly, reduced surface expression, and impaired function on neuronal membrane, causing neurological disorders including epilepsy and intellectual disability. Here, we investigated the proteostasis maintenance of NMDARs containing epilepsy-associated variations in the GluN2A subunit, including M705V and A727T. In the transfected HEK293T cells, we showed that the two variants were targeted to the proteasome for degradation and had reduced functional surface expression. We demonstrated that the application of BIX, a known small molecule activator of an HSP70 family chaperone BiP (binding immunoglobulin protein) in the endoplasmic reticulum (ER), dose-dependently enhanced the functional surface expression of the M705V and A727T variants in HEK293T cells. Moreover, BIX (10 μM) increased the surface protein levels of the M705V variant in human iPSC-derived neurons. We revealed that BIX promoted folding, inhibited degradation, and enhanced anterograde trafficking of the M705V variant by modest activation of the IRE1 pathway of the unfolded protein response. Our results suggest that adapting the ER proteostasis network restores the folding, trafficking, and function of pathogenic NMDAR variants, representing a potential treatment for neurological disorders resulting from NMDAR dysfunction.

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Fig. 1: The alternations as a result of variants in GluN2A subunit on the expression and function of trafficking-deficient NMDARs.
Fig. 2: The GluN2A variants are retained in the ER and degraded by the proteasome.
Fig. 3: Effects of BIX on the expression and function of trafficking-deficient variants in GluN2A subunit of NMDARs.
Fig. 4: Effects of BIX treatment on the surface expression of a trafficking-deficient GluN2A(M705V) variant in hiPSC-derived neurons.
Fig. 5: Effect of BIX treatment on the folding, assembly, trafficking, and degradation of variant NMDARs.
Fig. 6: Effect of BIX on increasing the GluN2A variant total protein level is not dependent on BiP.
Fig. 7: BIX increases variant GluN2A subunits expression through pharmacological activation of the unfolded protein response (UPR).
Fig. 8: Proposed mechanism of action of BIX effect on restoring proteostasis of NMDARs harboring GluN2A pathogenic variations.

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Acknowledgements

This work was supported by the National Institutes of Health (R01NS117176 to TWM, R01NS123524 to AES, and F30HD110088 to LYA) and the Brain Research Foundation (BRFSG-2021-08 to AES).

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Authors and Affiliations

  1. Department of Physiology and Biophysics, Case Western Reserve University School of Medicine, Cleveland, OH, 44106, USA

    Pei-pei Zhang, Taylor M. Benske, Ting-wei Mu & Ya-juan Wang

  2. Department of Genetics and Genome Sciences, Case Western Reserve University School of Medicine, Cleveland, OH, 44106, USA

    Lucie Y. Ahn & Ashleigh E. Schaffer

  3. Research Centre for Infectious Diseases, Department of Molecular and Biomedical Science, University of Adelaide, Adelaide, SA, 5005, Australia

    James C. Paton & Adrienne W. Paton

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  1. Pei-pei Zhang
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Contributions

PPZ, TWM, and YJW designed research; PPZ, LYA, and YJW performed research; PPZ, TWM, and YJW analyzed data; JCP and AWP contributed reagents; PPZ, TWM and YJW wrote the original draft; PPZ, TMB, LYA, AES, JCP, AWP, TWM, and YJW reviewed and edited the manuscript. All authors discussed the study.

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Correspondence to Ting-wei Mu or Ya-juan Wang.

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Zhang, Pp., Benske, T.M., Ahn, L.Y. et al. Adapting the endoplasmic reticulum proteostasis rescues epilepsy-associated NMDA receptor variants. Acta Pharmacol Sin 45, 282–297 (2024). https://doi.org/10.1038/s41401-023-01172-w

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