Newly-formed synaptic vesicles (SVs) are rapidly acidified by vacuolar adenosine triphosphatases (vATPases), generating a proton electrochemical gradient that drives neurotransmitter loading. Clathrin-mediated endocytosis is needed for the formation of new SVs, yet it is unclear when endocytosed vesicles acidify and refill at the synapse. Here, we isolated clathrin-coated vesicles (CCVs) from mouse brain to measure their acidification directly at the single vesicle level. We observed that the ATP-induced acidification of CCVs was strikingly reduced in comparison to SVs. Remarkably, when the coat was removed from CCVs, uncoated vesicles regained ATP-dependent acidification, demonstrating that CCVs contain the functional vATPase, yet its function is inhibited by the clathrin coat. Considering the known structures of the vATPase and clathrin coat, we propose a model in which the formation of the coat surrounds the vATPase and blocks its activity. Such inhibition is likely fundamental for the proper timing of SV refilling.
The structure has been deposited with the EMDB-ID #4335.For additional information considering structure please contact Prof Dr Carsten Mim at email@example.com
- Ira Milosevic
- Carsten Mim
- Ira Milosevic
- Zohreh Farsi
- Sindhuja Gowrisankaran
- Eileen M Lafer
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Animal experimentation: Animal experiments were conducted according to the European Guidelines for animal welfare (2010/63/EU) with approval by the Lower Saxony Landesamt fur Verbraucherschutz und Lebensmittelsicherheit (LAVES), registration number 14/1701.
- Margaret S. Robinson, University of Cambridge, United Kingdom
© 2018, Farsi et al.
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