Sec17/Sec18 can support membrane fusion without help from completion of SNARE zippering
- Geisel School of Medicine at Dartmouth, United States
- Stanford University, United States
Abstract
Membrane fusion requires R-, Qa-, Qb-, and Qc-family SNAREs that zipper into RQaQbQc coiled coils, driven by the sequestration of apolar amino acids. Zippering has been thought to provide all the force driving fusion. Sec17/aSNAP can form an oligomeric assembly with SNAREs with the Sec17 C-terminus bound to Sec18/NSF, the central region bound to SNAREs, and a crucial apolar loop near the N-terminus poised to insert into membranes. We now report that Sec17 and Sec18 will drive robust fusion without requiring zippering completion. Zippering-driven fusion is blocked by deleting the C-terminal quarter of any Q-SNARE domain or by replacing the apolar amino acids of the Qa-SNARE which face the center of the 4-SNARE coiled coils with polar residues. These blocks, singly or combined, are bypassed by Sec17 and Sec18, and SNARE-dependent fusion is restored without help from completing zippering.
Data availability
All data generated or analyzed during this study are included in the manuscript and supporting files. Source data files have been provided for Figures 1, 2, 3 4, 5, and 6.
Article and author information
Author details
Funding
National Institutes of Health (R35GM118037)
- William T Wickner
National Institutes of Health (R37MH63105)
- Axel T Brunger
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Josep Rizo, University of Texas Southwestern Medical Center, United States
Version history
- Received: February 16, 2021
- Accepted: April 30, 2021
- Accepted Manuscript published: May 4, 2021 (version 1)
- Version of Record published: May 24, 2021 (version 2)
Copyright
© 2021, Song et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Sec17/Sec18 can support membrane fusion without help from completion of SNARE zippering
eLife 10:e67578.
https://doi.org/10.7554/eLife.67578
Further reading
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- Biochemistry and Chemical Biology
Two proteins called Sec17 and Sec18 may have a larger role in membrane fusion than is commonly assumed in textbook models.
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- Biochemistry and Chemical Biology
- Plant Biology
Metabolism and biological functions of the nitrogen-rich compound guanidine have long been neglected. The discovery of four classes of guanidine-sensing riboswitches and two pathways for guanidine degradation in bacteria hint at widespread sources of unconjugated guanidine in nature. So far, only three enzymes from a narrow range of bacteria and fungi have been shown to produce guanidine, with the ethylene-forming enzyme (EFE) as the most prominent example. Here, we show that a related class of Fe2+- and 2-oxoglutarate-dependent dioxygenases (2-ODD-C23) highly conserved among plants and algae catalyze the hydroxylation of homoarginine at the C6-position. Spontaneous decay of 6-hydroxyhomoarginine yields guanidine and 2-aminoadipate-6-semialdehyde. The latter can be reduced to pipecolate by pyrroline-5-carboxylate reductase but more likely is oxidized to aminoadipate by aldehyde dehydrogenase ALDH7B in vivo. Arabidopsis has three 2-ODD-C23 isoforms, among which Din11 is unusual because it also accepted arginine as substrate, which was not the case for the other 2-ODD-C23 isoforms from Arabidopsis or other plants. In contrast to EFE, none of the three Arabidopsis enzymes produced ethylene. Guanidine contents were typically between 10 and 20 nmol*(g fresh weight)-1 in Arabidopsis but increased to 100 or 300 nmol*(g fresh weight)-1 after homoarginine feeding or treatment with Din11-inducing methyljasmonate, respectively. In 2-ODD-C23 triple mutants, the guanidine content was strongly reduced, whereas it increased in overexpression plants. We discuss the implications of the finding of widespread guanidine-producing enzymes in photosynthetic eukaryotes as a so far underestimated branch of the bio-geochemical nitrogen cycle and propose possible functions of natural guanidine production.
