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Mol Biol Cell. 2013 Apr;24(8):1176-84. doi: 10.1091/mbc.E12-11-0810. Epub 2013 Feb 20.

Doc2b promotes GLUT4 exocytosis by activating the SNARE-mediated fusion reaction in a calcium- and membrane bending-dependent manner.

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1
Department of Molecular, Cellular and Developmental Biology, University of Colorado at Boulder, Boulder, CO 80309, USA.

Abstract

The glucose transporter GLUT4 plays a central role in maintaining body glucose homeostasis. On insulin stimulation, GLUT4-containing vesicles fuse with the plasma membrane, relocating GLUT4 from intracellular reservoirs to the cell surface to uptake excess blood glucose. The GLUT4 vesicle fusion reaction requires soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) as the core fusion engine and a group of regulatory proteins. In particular, the soluble C2-domain factor Doc2b plays a key role in GLUT4 vesicle fusion, but its molecular mechanism has been unclear. Here we reconstituted the SNARE-dependent GLUT4 vesicle fusion in a defined proteoliposome fusion system. We observed that Doc2b binds to GLUT4 exocytic SNAREs and potently accelerates the fusion kinetics in the presence of Ca(2+). The stimulatory activity of Doc2b requires intact Ca(2+)-binding sites on both the C2A and C2B domains. Using electron microscopy, we observed that Doc2b strongly bends the membrane bilayer, and this membrane-bending activity is essential to the stimulatory function of Doc2b in fusion. These results demonstrate that Doc2b promotes GLUT4 exocytosis by accelerating the SNARE-dependent fusion reaction by a Ca(2+)- and membrane bending-dependent mechanism. Of importance, certain features of Doc2b function appear to be distinct from how synaptotagmin-1 promotes synaptic neurotransmitter release, suggesting that exocytic Ca(2+) sensors may possess divergent mechanisms in regulating vesicle fusion.

PMID:
23427263
PMCID:
PMC3623638
DOI:
10.1091/mbc.E12-11-0810
[Indexed for MEDLINE]
Free PMC Article
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