RT Journal Article
SR Electronic
T1 Nanoscaled RIM clustering at presynaptic active zones revealed by endogenous tagging
JF Life Science Alliance
JO Life Sci. Alliance
FD Life Science Alliance LLC
SP e202302021
DO 10.26508/lsa.202302021
VO 6
IS 12
A1 Mrestani, Achmed
A1 Dannhäuser, Sven
A1 Pauli, Martin
A1 Kollmannsberger, Philip
A1 Hübsch, Martha
A1 Morris, Lydia
A1 Langenhan, Tobias
A1 Heckmann, Manfred
A1 Paul, Mila M
YR 2023
UL http://www.life-science-alliance.org/content/6/12/e202302021.abstract
AB Chemical synaptic transmission involves neurotransmitter release from presynaptic active zones (AZs). The AZ protein Rab-3-interacting molecule (RIM) is important for normal Ca2+-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 rimV5−Znf but not in rimHA−Znf. In addition, rimHA−Znf synapses show intact presynaptic homeostatic potentiation. Combining super-resolution localization microscopy and hierarchical clustering, we detect ∼10 RIMHA−Znf subclusters with ∼13 nm diameter per AZ that are compacted and increased in numbers in presynaptic homeostatic potentiation.