Li, Feng;
Sundaram, Ramalingam Venkat Kalyana;
Gatta, Alberto T;
Coleman, Jeff;
Ramakrishnan, Sathish;
Krishnakumar, Shyam S;
Pincet, Frederic;
(2021)
Vesicle capture by membrane-bound Munc13-1 requires self-assembly into discrete clusters.
FEBS Letters
, 595
(17)
pp. 2185-2196.
10.1002/1873-3468.14157.
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Abstract
Munc13-1 is a large banana-shaped soluble protein that is involved in the regulation of synaptic vesicle docking and fusion. Recent studies suggest that multiple copies of Munc13-1 form nano-assemblies in active zones of neurons. However, it is not known whether such clustering of Munc13-1 is correlated with multivalent binding to synaptic vesicles or specific plasma membrane domains at docking sites in the active zone. The functional significance of putative Munc13-1 clustering is also unknown. Here, we report that nano-clustering is an inherent property of Munc13-1 and is indeed required for vesicle binding to bilayers containing Munc13-1. Purified Munc13-1 protein reconstituted onto supported lipid bilayers assembled into clusters containing from 2 to ˜ 20 copies as revealed by a combination of quantitative TIRF microscopy and step-wise photobleaching. Surprisingly, only clusters containing a minimum of 6 copies of Munc13-1 were capable of efficiently capturing and retaining small unilamellar vesicles. The C-terminal C2C domain of Munc13-1 is not required for Munc13-1 clustering, but is required for efficient vesicle capture. This capture is largely due to a combination of electrostatic and hydrophobic interactions between the C2C domain and the vesicle membrane.
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