Molecular in situ topology of Aczonin/Piccolo and associated proteins at the mammalian neurotransmitter release site
Open accessAbstract
The protein machinery of neurotransmitter exocytosis requires efficient orchestration in space and time, for speed and precision of neurotransmission and also for synaptic ontogeny and plasticity. However, its spatial organization in situ is virtually unknown. Aczonin/Piccolo is a putative organizer protein of mammalian active zones. We determined by immunogold electron microscopy (EM) ( i ) the spatial arrangement (i.e., topology) of 11 segments of the Aczonin polypeptide in situ, and correlated it to ( ii ) the positioning of Aczonin-interacting domains of Bassoon, CAST/ELKS, Munc13, and RIM and ( iii ) the ultrastructurally defined presynaptic macromolecular aggregates known as dense projections and synaptic ribbons. At conventional synapses, Aczonin assumes a compact molecular topology within a layer 35 to 80 nm parallel to the plasma membrane (PM), with a “trunk” sitting on the dense projection top and a C-terminal “arm” extending down toward the PM and sideward to the dense projection periphery. At ribbon synapses, Aczonin occupies the whole ribbon area. Bassoon colocalizes with Aczonin at conventional synapses but not at ribbon synapses. At both conventional and ribbon synapses, CAST, Munc13, and RIM are segregated from Aczonin, closer to the PM, and Aczonin is positioned such that it may control the access of neurotransmitter vesicles to the fusion site.
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EM studies of chemically fixed and stained central synapses showed that the active zone contains a hexagonal grid of dense projections with intercalated vesicles.
“EM studies of chemically fixed and stained central synapses showed that the active zone contains a hexagonal grid of dense projections with intercalated vesicles (Figure 4A; Akert et al., 1972; Pfenninger et al., 1972; Limbach et al., 2011).”
From The Presynaptic Active Zone (2012) · cites this paper as Limbach et al., 2011
The introduction (page unavailable) says that EM of heavy-metal-stained mammalian synapses reveals regularly spaced dense projections forming a “presynaptic grid.” The “Dense Projections: Fact or Artifact?” section describes these structures in chemically fixed specimens and refers to docked vesicles within the grid, but our copy does not specify a hexagonal arrangement or say the vesicles are intercalated. (LLM, conf=0.85)
Experimental — accuracy not yet measuredRIM and Munc13 are localized between the dense projections adjacent to the plasma membrane, whereas piccolo and bassoon are more distant and appear to be attached to the tips of the dense projections.
“Immuno-EM experiments suggested that RIM and Munc13, arguably the two most important active zone proteins, are localized between the dense projections adjacent to the plasma membrane, whereas the cytomatrix proteins piccolo and bassoon are more distant and appear to be attached to the tips of the dense projections (Limbach et al., 2011).”
From The Presynaptic Active Zone (2012) · cites this paper as Limbach et al., 2011
Munc13-1 in synapses appears tightly anchored to the active zone via scaffold proteins and interacts with components of the sub-membranous cytoskeleton, rendering it effectively insoluble.
“Although Munc13s do not have transmembrane domains, Munc13-1 in synapses appears to be tightly anchored to the AZ via scaffold proteins [62,63] and interacts with components of the sub-membranous cytoskeleton [64], rendering it effectively insoluble [65].”
From Presynaptic cAMP-PKA-mediated potentiation induces reconfiguration of synaptic vesicle pools and channel-vesicle coupling at hippocampal mossy fiber boutons (2024) · cites this paper as [63]
Citation [65] is attached specifically to the claim that Munc13-1 is effectively insoluble. The Spatial Relationships section describes Munc13’s location and possible movement between the membrane and cytosol, but the passages found by searching for insolubility and detergent extraction do not address whether it is insoluble. (LLM, conf=0.75)
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Cites
The organization of cytoplasm at the presynaptic active zone of a central nervous system synapse
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Piccolo, a presynaptic zinc finger protein structurally related to bassoon
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Quantitative analysis of the native presynaptic cytomatrix by cryoelectron tomography
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Bassoon speeds vesicle reloading at a central excitatory synapse
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- Authors
- Limbach, C., Laue, M. M., Wang, X., Hu, B., Thiede, N., Hultqvist, G., & Kilimann, M. W.
- Affiliations
- Uppsala University (SE)
- Ruhr University Bochum (DE)
- Corresponding author
- Kilimann, Manfred W.
- Venue
- Proceedings of the National Academy of Sciences
- Publisher
- National Academy of Sciences
- Published
- 2011-06-28
- Volume
- 108
- Issue
- 31
- Pages
- E392-401
- ISSN
- 0027-8424 (ISSN-L)
- 1091-6490
- PMC
- PMC3150911
- OpenAlex
- W2011381493
- Cited by
- 92
- References
- 5
- Citations by year
- 2026: 6 · 2025: 4 · 2024: 7 · 2023: 3 · 2022: 7 · 2021: 5 · 2020: 4 · 2019: 7 · 2018: 5 · 2017: 10 · 2016: 4 · 2015: 10 · 2014: 9 · 2013: 5 · 2012: 5
- FWCI
- 2.65
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Locations
- Proceedings of the National Academy of Sciences (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Publication server of the Robert Koch Institute (Robert Kock Institute) (submittedVersion)
- Funders
- European Commission
- Deutsche Forschungsgemeinschaft
- Uppsala Universitet
- Topics
- Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Neuroscience and Neuropharmacology Research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular transport and secretion
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Extracellular vesicles in disease
- Keywords
- piccolo, active zone, neurotransmitter release, synaptic vesicle exocytosis, ribbon synapses, bassoon, Munc13, RIM, synaptic ribbons, immunogold electron microscopy
- MeSH terms
- Animals, Binding Sites, Cell Membrane, Cytoskeletal Proteins, Nerve Tissue Proteins, Neuropeptides, Protein Binding, Synapses, Immunoblotting, Microscopy, Immunoelectron, Rats, Sprague-Dawley, Neurotransmitter Agents, GTP-Binding Proteins, Multiprotein Complexes, Adaptor Proteins, Signal Transducing, Rats
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