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Munc13 C2B domain is an activity-dependent Ca2+ regulator of synaptic exocytosis

Nature Structural & Molecular Biology volume 17, pages 280–288 (2010)Cite this article

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Abstract

Munc13 is a multidomain protein present in presynaptic active zones that mediates the priming and plasticity of synaptic vesicle exocytosis, but the mechanisms involved remain unclear. Here we use biophysical, biochemical and electrophysiological approaches to show that the central C2B domain of Munc13 functions as a Ca2+ regulator of short-term synaptic plasticity. The crystal structure of the C2B domain revealed an unusual Ca2+-binding site with an amphipathic α-helix. This configuration confers onto the C2B domain unique Ca2+-dependent phospholipid-binding properties that favor phosphatidylinositolphosphates. A mutation that inactivated Ca2+-dependent phospholipid binding to the C2B domain did not alter neurotransmitter release evoked by isolated action potentials, but it did depress release evoked by action-potential trains. In contrast, a mutation that increased Ca2+-dependent phosphatidylinositolbisphosphate binding to the C2B domain enhanced release evoked by isolated action potentials and by action-potential trains. Our data suggest that, during repeated action potentials, Ca2+ and phosphatidylinositolphosphate binding to the Munc13 C2B domain potentiate synaptic vesicle exocytosis, thereby offsetting synaptic depression induced by vesicle depletion.

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Figure 1: The Munc13 C2B domain is a Ca2+-binding module.
Figure 2: Three-dimensional structures of the Ca2+-free and Ca2+-bound Munc13-1 C2B domain.
Figure 3: Ca2+-dependent binding of the Munc13 (M13) C2B domain to PIP- or PIP2-containing liposomes.
Figure 4: PIP and PIP2 dependence of Ca2+-induced liposome binding to Munc13 C2B domains.
Figure 5: Effect of Munc13-2 C2B domain mutations on release induced by isolated action potentials.
Figure 6: Ca2+ binding to the Munc13 C2B domain regulates release during high-frequency action-potential trains.
Figure 7: Model for the Ca2+ regulation of short-term plasticity by Munc13.

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Acknowledgements

We thank I. Kornblum, I. Herfort and H. Deng for excellent technical support. This paper was supported by grants from the US National Institutes of Health (NS051262 to C.R.; NS40944 to J.R.) and the Deutsche Forschungsgemeinschat (to C.R.).

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Author notes

  1. Ok-Ho Shin, Jun Lu & Jeong-Seop Rhee

    Present address: Present addresses: Department of Neuroscience & Cell Biology, University of Texas Medical Branch, Galveston, Texas, USA (O.-H.S.); Merck & Co. Inc., Rahway, New Jersey, USA (J.L.); Department of Molecular Neurobiology, Max-Planck-Institut für Experimentelle Medizin, Göttingen, Germany (J.-S.R.).,

  2. Ok-Ho Shin, Jun Lu and Jeong-Seop Rhee: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Neuroscience, University of Texas Southwestern Medical Center, Dallas, Texas, USA

    Ok-Ho Shin, Zhiping P Pang & Thomas C Südhof

  2. Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, Texas, USA

    Jun Lu, Diana R Tomchick, Mischa Machius & Josep Rizo

  3. Department of Membrane Biophysics, Max-Planck-Institut für Biophysikalische Chemie, Göttingen, Germany

    Jeong-Seop Rhee & Christian Rosenmund

  4. Department of Neuroscience, Baylor College of Medicine, Houston, Texas, USA

    Jeong-Seop Rhee, Marcial Camacho-Perez & Christian Rosenmund

  5. Department of Molecular & Cellular Physiology, Stanford University, California, USA

    Zhiping P Pang & Thomas C Südhof

  6. Department of Molecular Neurobiology, Max-Planck-Institut für Experimentelle Medizin, Göttingen, Germany

    Sonja M Wojcik & Nils Brose

  7. Department of Molecular & Human Genetics, Baylor College of Medicine, Houston, Texas, USA

    Marcial Camacho-Perez

  8. Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, Texas, USA

    Josep Rizo & Christian Rosenmund

  9. Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, Texas, USA

    Thomas C Südhof

  10. Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, Texas, USA

    Thomas C Südhof

  11. Howard Hughes Medical Institute, Stanford University, Stanford, California, USA

    Thomas C Südhof

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Contributions

O.-H. S. performed the protein chemistry and molecular biology experiments; J.L., D.R.T. and M.M. performed the structural biology experiments; J.-S.R., M.C.-P. and Z.P.P. performed the electrophysiology experiments; S.W. and N.B. generated the vectors for expression of mutant Munc13; J.R., C.R. and T.C.S. wrote the paper.

Corresponding authors

Correspondence to Josep Rizo, Christian Rosenmund or Thomas C Südhof.

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Shin, OH., Lu, J., Rhee, JS. et al. Munc13 C2B domain is an activity-dependent Ca2+ regulator of synaptic exocytosis. Nat Struct Mol Biol 17, 280–288 (2010). https://doi.org/10.1038/nsmb.1758

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