title: An acto-myosin II constricting ring initiates the fission of activity-dependent bulk endosomes in neurosecretory cells.
abstract: Activity-dependent bulk endocytosis allows neurons to internalize large portions of the plasma membrane in response to stimulation. However, whether this critical type of compensatory endocytosis is unique to neurons or also occurs in other excitable cells is currently unknown. Here we used fluorescent 70 kDa dextran to demonstrate that secretagogue-induced bulk endocytosis also occurs in bovine chromaffin cells. The relatively large size of the bulk endosomes found in this model allowed us to investigate how the neck of the budding endosomes constricts to allow efficient recruitment of the fission machinery. Using time-lapse imaging of Lifeact-GFP-transfected chromaffin cells in combination with fluorescent 70 kDa dextran, we detected acto-myosin II rings surrounding dextran-positive budding endosomes. Importantly, these rings were transient and contracted before disappearing, suggesting that they might be involved in restricting the size of the budding endosome neck. Based on the complete recovery of dextran fluorescence after photobleaching, we demonstrated that the actin ring-associated budding endosomes were still connected with the extracellular fluid. In contrast, no such recovery was observed following the constriction and disappearance of the actin rings, suggesting that these structures were pinched-off endosomes. Finally, we showed that the rings were initiated by a circular array of phosphatidylinositol(4,5)bisphosphate microdomains, and that their constriction was sensitive to both myosin II and dynamin inhibition. The acto-myosin II rings therefore play a key role in constricting the neck of budding bulk endosomes before dynamin-dependent fission from the plasma membrane of neurosecretory cells.
authors: The Clem Jones Centre for Ageing Dementia Research, Queensland Brain Institute, University of Queensland, Brisbane 4072, Queensland, Australia.
authors: Rachel S
authors: RS
authors: Gormal
authors: The Clem Jones Centre for Ageing Dementia Research, Queensland Brain Institute, University of Queensland, Brisbane 4072, Queensland, Australia.
authors: Tam H
authors: TH
authors: Nguyen
authors: The Clem Jones Centre for Ageing Dementia Research, Queensland Brain Institute, University of Queensland, Brisbane 4072, Queensland, Australia.
authors: Sally
authors: http://orcid.org/0000-0001-9294-5404
authors: S
authors: Martin
authors: The Clem Jones Centre for Ageing Dementia Research, Queensland Brain Institute, University of Queensland, Brisbane 4072, Queensland, Australia.
authors: Andreas
authors: A
authors: Papadopulos
authors: The Clem Jones Centre for Ageing Dementia Research, Queensland Brain Institute, University of Queensland, Brisbane 4072, Queensland, Australia f.meunier@uq.edu.au.
authors: Frederic A
authors: FA
authors: Meunier
mesh_terms: D000199:Actins / Q000378:metabolism*; D000311:Adrenal Glands / Q000166:cytology; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D019439:Chromaffin Cells / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure*; D003911:Dextrans / Q000378:metabolism; D034281:Dynamins / Q000037:antagonists & inhibitors; D004705:Endocytosis / Q000187:drug effects / Q000502:physiology*; D011992:Endosomes / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D017261:Glycosylphosphatidylinositols / Q000235:genetics / Q000378:metabolism; D006576:Heterocyclic Compounds, 4 or More Rings / Q000494:pharmacology; D006835:Hydrazones / Q000494:pharmacology; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D024462:Myosin Type II / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009284:Naphthols / Q000494:pharmacology; D009538:Nicotine / Q000494:pharmacology; D018722:Nicotinic Agonists / Q000494:pharmacology; D012235:Rhodamines / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
chemical_list: D000199:Actins; D003911:Dextrans; C067661:Fluoro-Ruby; D017261:Glycosylphosphatidylinositols; D006576:Heterocyclic Compounds, 4 or More Rings; D006835:Hydrazones; D008164:Luminescent Proteins; D009284:Naphthols; D018722:Nicotinic Agonists; D012235:Rhodamines; C568360:dyngo-4a; C472645:blebbistatin; D009538:Nicotine; D024462:Myosin Type II; D034281:Dynamins
keywords: PIP2; actin; cytoskeleton; dynamin; endocytosis; myosin II
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