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Nanoscaled RIM clustering at presynaptic active zones revealed by endogenous tagging

Chemical synaptic transmission involves neurotransmitter release from presynaptic active zones (AZs). The AZ protein Rab-3-interacting molecule (RIM) is important for normal Ca(2+)-triggered release. However, its precise localization within AZs of the glutamatergic neuromuscular junctions of Drosoph...

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Autores principales: Mrestani, Achmed, Dannhäuser, Sven, Pauli, Martin, Kollmannsberger, Philip, Hübsch, Martha, Morris, Lydia, Langenhan, Tobias, Heckmann, Manfred, Paul, Mila M
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Life Science Alliance LLC 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10494931/
https://www.ncbi.nlm.nih.gov/pubmed/37696575
http://dx.doi.org/10.26508/lsa.202302021
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author Mrestani, Achmed
Dannhäuser, Sven
Pauli, Martin
Kollmannsberger, Philip
Hübsch, Martha
Morris, Lydia
Langenhan, Tobias
Heckmann, Manfred
Paul, Mila M
author_facet Mrestani, Achmed
Dannhäuser, Sven
Pauli, Martin
Kollmannsberger, Philip
Hübsch, Martha
Morris, Lydia
Langenhan, Tobias
Heckmann, Manfred
Paul, Mila M
author_sort Mrestani, Achmed
collection PubMed
description Chemical synaptic transmission involves neurotransmitter release from presynaptic active zones (AZs). The AZ protein Rab-3-interacting molecule (RIM) is important for normal Ca(2+)-triggered release. However, its precise localization within AZs of the glutamatergic neuromuscular junctions of Drosophila melanogaster remains elusive. We used CRISPR/Cas9-assisted genome engineering of the rim locus to incorporate small epitope tags for targeted super-resolution imaging. A V5-tag, derived from simian virus 5, and an HA-tag, derived from human influenza virus, were N-terminally fused to the RIM Zinc finger. Whereas both variants are expressed in co-localization with the core AZ scaffold Bruchpilot, electrophysiological characterization reveals that AP-evoked synaptic release is disturbed in rim(V5−Znf) but not in rim(HA−Znf). In addition, rim(HA−Znf) synapses show intact presynaptic homeostatic potentiation. Combining super-resolution localization microscopy and hierarchical clustering, we detect ∼10 RIM(HA−Znf) subclusters with ∼13 nm diameter per AZ that are compacted and increased in numbers in presynaptic homeostatic potentiation.
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spelling pubmed-104949312023-09-12 Nanoscaled RIM clustering at presynaptic active zones revealed by endogenous tagging Mrestani, Achmed Dannhäuser, Sven Pauli, Martin Kollmannsberger, Philip Hübsch, Martha Morris, Lydia Langenhan, Tobias Heckmann, Manfred Paul, Mila M Life Sci Alliance Research Articles Chemical synaptic transmission involves neurotransmitter release from presynaptic active zones (AZs). The AZ protein Rab-3-interacting molecule (RIM) is important for normal Ca(2+)-triggered release. However, its precise localization within AZs of the glutamatergic neuromuscular junctions of Drosophila melanogaster remains elusive. We used CRISPR/Cas9-assisted genome engineering of the rim locus to incorporate small epitope tags for targeted super-resolution imaging. A V5-tag, derived from simian virus 5, and an HA-tag, derived from human influenza virus, were N-terminally fused to the RIM Zinc finger. Whereas both variants are expressed in co-localization with the core AZ scaffold Bruchpilot, electrophysiological characterization reveals that AP-evoked synaptic release is disturbed in rim(V5−Znf) but not in rim(HA−Znf). In addition, rim(HA−Znf) synapses show intact presynaptic homeostatic potentiation. Combining super-resolution localization microscopy and hierarchical clustering, we detect ∼10 RIM(HA−Znf) subclusters with ∼13 nm diameter per AZ that are compacted and increased in numbers in presynaptic homeostatic potentiation. Life Science Alliance LLC 2023-09-11 /pmc/articles/PMC10494931/ /pubmed/37696575 http://dx.doi.org/10.26508/lsa.202302021 Text en © 2023 Mrestani et al. https://creativecommons.org/licenses/by/4.0/This article is available under a Creative Commons License (Attribution 4.0 International, as described at https://creativecommons.org/licenses/by/4.0/).
spellingShingle Research Articles
Mrestani, Achmed
Dannhäuser, Sven
Pauli, Martin
Kollmannsberger, Philip
Hübsch, Martha
Morris, Lydia
Langenhan, Tobias
Heckmann, Manfred
Paul, Mila M
Nanoscaled RIM clustering at presynaptic active zones revealed by endogenous tagging
title Nanoscaled RIM clustering at presynaptic active zones revealed by endogenous tagging
title_full Nanoscaled RIM clustering at presynaptic active zones revealed by endogenous tagging
title_fullStr Nanoscaled RIM clustering at presynaptic active zones revealed by endogenous tagging
title_full_unstemmed Nanoscaled RIM clustering at presynaptic active zones revealed by endogenous tagging
title_short Nanoscaled RIM clustering at presynaptic active zones revealed by endogenous tagging
title_sort nanoscaled rim clustering at presynaptic active zones revealed by endogenous tagging
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10494931/
https://www.ncbi.nlm.nih.gov/pubmed/37696575
http://dx.doi.org/10.26508/lsa.202302021
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