@article{ba1259b21e4140cab58879326111cfdd,
title = "Presynaptic targeting of botulinum neurotoxin type A requires a tripartite PSG-Syt1-SV2 plasma membrane nanocluster for synaptic vesicle entry",
abstract = "The unique nerve terminal targeting of botulinum neurotoxin type A (BoNT/A) is due to its capacity to bind two receptors on the neuronal plasma membrane: polysialoganglioside (PSG) and synaptic vesicle glycoprotein 2 (SV2). Whether and how PSGs and SV2 may coordinate other proteins for BoNT/A recruitment and internalization remains unknown. Here, we demonstrate that the targeted endocytosis of BoNT/A into synaptic vesicles (SVs) requires a tripartite surface nanocluster. Live-cell super-resolution imaging and electron microscopy of catalytically inactivated BoNT/A wildtype and receptor-binding-deficient mutants in cultured hippocampal neurons demonstrated that BoNT/A must bind coincidentally to a PSG and SV2 to target synaptic vesicles. We reveal that BoNT/A simultaneously interacts with a preassembled PSG-synaptotagmin-1 (Syt1) complex and SV2 on the neuronal plasma membrane, facilitating Syt1-SV2 nanoclustering that controls endocytic sorting of the toxin into synaptic vesicles. Syt1 CRISPRi knockdown suppressed BoNT/A- and BoNT/E-induced neurointoxication as quantified by SNAP-25 cleavage, suggesting that this tripartite nanocluster may be a unifying entry point for selected botulinum neurotoxins that hijack this for synaptic vesicle targeting.",
author = "Merja Joensuu and Parnayan Syed and Saber, {Saber H} and Vanessa Lanoue and Wallis, {Tristan P} and James Rae and Ailisa Blum and Gormal, {Rachel S} and Christopher Small and Shanley Sanders and Anmin Jiang and Stefan Mahrhold and Nadja Krez and Cousin, {Michael A} and Ruby Cooper-White and Cooper-White, {Justin J} and Collins, {Brett M} and Parton, {Robert G} and Giuseppe Balistreri and Andreas Rummel and Meunier, {Frederic A}",
note = "Funding Information: We would like to thank the DNA Dream Lab (DDL) facility (HiLIFE, Institute of Biotechnology, Helsinki Finland) and Konstantin Kogan for the design and construction of the oligonucleotides listed in Table EV2 and DNA plasmids listed in Table EV3 . We would like to acknowledge the facilities, and the scientific and technical assistance, of the Microscopy Australia Facility at the Centre for Microscopy and Microanalysis (CMM), especially Dr Kathryn Green, and the Queensland Brain Institute (QBI) Advanced Microscopy Facility, and the facility staff, especially Dr Rumelo Amor, at The University of Queensland. We would like to extend our thanks to Prof Claudio Rivera Baeza (Neuroscience Center, Helsinki Institute of Life Science, University of Helsinki, Finland), Drs Mariana Nelson, Ron Broide, and Birgitte Jacky (Allergan/Abbvie) for insightful discussions, Dr Alex McCann (QBI) for her help with the manuscript formatting and Rowan Tweedale (QBI) for critical appraisal of the manuscript. We would also like to thank Thomas Binz (Medizinische Hochschule Hannover, Germany) for kindly providing BoNT/E, and D. Sesardic and P. Stickings (Division of Bacteriology, National Institute for Biological Standards and Control, United Kingdom) for providing SNAP‐25/A and SNAP‐25/E antibodies. This work is supported by the Australian Research Council (ARC) Discovery Early Career Researcher Award (DE190100565) and The University of Queensland Amplify fellowship to MJ, ARC LIEF grant (LE130100078), National Health and Medical Research Council of Australia (NHMRC) Project Grant (1120381) and NHMRC Senior Research Fellowship (1155794) to FAM, NHMRC Project Grants (1140064 and 1150083), and Fellowship (1156489) to RGP, NHMRC Senior Research Fellowship (1136021) to BMC, Academy of Finland Grant (318434) to GB and German Federal Ministry of Education and Research grants 031L0111B (TiViBoNT) and 13N15512 (X‐BAT) to AR. Open access publishing facilitated by The University of Queensland, as part of the Wiley ‐ The University of Queensland agreement via the Council of Australian University Librarians. wt Publisher Copyright: {\textcopyright} 2023 The Authors. Published under the terms of the CC BY 4.0 license.",
year = "2023",
month = may,
day = "25",
doi = "10.15252/embj.2022112095",
language = "English",
journal = "EMBO Journal",
issn = "0261-4189",
publisher = "Wiley-Blackwell",
}