In nerve cells the genes encoding for α2δ subunits of voltage-gated calcium channels have been linked to synaptic functions and neurological disease. Here we show that α2δ subunits are essential for the formation and organization of glutamatergic synapses. Using a cellular α2δ subunit triple-knockout/knockdown model, we demonstrate a failure in presynaptic differentiation evidenced by defective presynaptic calcium channel clustering and calcium influx, smaller presynaptic active zones, and a strongly reduced accumulation of presynaptic vesicle-associated proteins (synapsin and vGLUT). The presynaptic defect is associated with the downscaling of postsynaptic AMPA receptors and the postsynaptic density. The role of α2δ isoforms as synaptic organizers is highly redundant, as each individual α2δ isoform can rescue presynaptic calcium channel trafficking and expression of synaptic proteins. Moreover, α2δ-2 and α2δ-3 with mutated metal ion-dependent adhesion sites can fully rescue presynaptic synapsin expression but only partially calcium channel trafficking, suggesting that the regulatory role of α2δ subunits is independent from its role as a calcium channel subunit. Our findings influence the current view on excitatory synapse formation. First, our study suggests that postsynaptic differentiation is secondary to presynaptic differentiation. Second, the dependence of presynaptic differentiation on α2δ implicates α2δ subunits as potential nucleation points for the organization of synapses. Finally, our results suggest that α2δ subunits act as transsynaptic organizers of glutamatergic synapses, thereby aligning the synaptic active zone with the postsynaptic density.
We thank Arnold Schwartz for providing α2δ-1 knockout mice; Ariane Benedetti, Sabine Baumgartner, Sandra Demetz, and Irene Mahlknecht for technical support; Nadine Ortner and Andreas Lieb for electrophysiological experiments; the team of the Electron Microscopy Facility at the Institute of Science and Technology Austria for technical support related to ultrastructural analysis; Hermann Dietrich and Anja Beierfuß and her team for animal care; Jutta Engel and Jörg Striessnig for critical discussions; and Bruno Benedetti and Bernhard Flucher for critical discussions and reading the manuscript. This study was supported by Austrian Science Fund Grants P24079, F44060, F44150, and DOC30-B30 (to G.J.O.) and T855 (to M.C.), European Research Council Grant AdG 694539 (to R.S.), Deutsche Forschungsgemeinschaft Grant SFB1348-TP A03 (to M.M.), and Interdisziplinäre Zentrum für Klinische Forschung Münster Grant Mi3/004/19 (to M.M.). This work is part of the PhD theses of C.L.S., S.M.G., and C.A.
Schöpf CL, Ablinger C, Geisler SM, et al. Presynaptic α2δ subunits are key organizers of glutamatergic synapses. PNAS. 2021;118(14). doi:10.1073/pnas.1920827118
Schöpf, C. L., Ablinger, C., Geisler, S. M., Stanika, R. I., Campiglio, M., Kaufmann, W., … Obermair, G. J. (2021). Presynaptic α2δ subunits are key organizers of glutamatergic synapses. PNAS. National Academy of Sciences. https://doi.org/10.1073/pnas.1920827118
Schöpf, Clemens L., Cornelia Ablinger, Stefanie M. Geisler, Ruslan I. Stanika, Marta Campiglio, Walter Kaufmann, Benedikt Nimmervoll, et al. “Presynaptic Α2δ Subunits Are Key Organizers of Glutamatergic Synapses.” PNAS. National Academy of Sciences, 2021. https://doi.org/10.1073/pnas.1920827118.
C. L. Schöpf et al., “Presynaptic α2δ subunits are key organizers of glutamatergic synapses,” PNAS, vol. 118, no. 14. National Academy of Sciences, 2021.
Schöpf CL, Ablinger C, Geisler SM, Stanika RI, Campiglio M, Kaufmann W, Nimmervoll B, Schlick B, Brockhaus J, Missler M, Shigemoto R, Obermair GJ. 2021. Presynaptic α2δ subunits are key organizers of glutamatergic synapses. PNAS. 118(14).
Schöpf, Clemens L., et al. “Presynaptic Α2δ Subunits Are Key Organizers of Glutamatergic Synapses.” PNAS, vol. 118, no. 14, National Academy of Sciences, 2021, doi:10.1073/pnas.1920827118.
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