A Central Small Amino Acid in the VAMP2 Transmembrane Domain Regulates the Fusion Pore in Exocytosis
Abstract Exocytosis depends on cytosolic domains of SNARE proteins but the function of the transmembrane domains (TMDs) in membrane fusion remains controversial. The TMD of the SNARE protein synaptobrevin2/VAMP2 contains two highly conserved small amino acids, G100 and C103, in its central portion....
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Nature Portfolio
2017
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oai:doaj.org-article:01d582e4c0cb4700b6227323642fc2002021-12-02T16:06:42ZA Central Small Amino Acid in the VAMP2 Transmembrane Domain Regulates the Fusion Pore in Exocytosis10.1038/s41598-017-03013-32045-2322https://doaj.org/article/01d582e4c0cb4700b6227323642fc2002017-06-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-03013-3https://doaj.org/toc/2045-2322Abstract Exocytosis depends on cytosolic domains of SNARE proteins but the function of the transmembrane domains (TMDs) in membrane fusion remains controversial. The TMD of the SNARE protein synaptobrevin2/VAMP2 contains two highly conserved small amino acids, G100 and C103, in its central portion. Substituting G100 and/or C103 with the β-branched amino acid valine impairs the structural flexibility of the TMD in terms of α-helix/β-sheet transitions in model membranes (measured by infrared reflection-absorption or evanescent wave spectroscopy) during increase in protein/lipid ratios, a parameter expected to be altered by recruitment of SNAREs at fusion sites. This structural change is accompanied by reduced membrane fluidity (measured by infrared ellipsometry). The G100V/C103V mutation nearly abolishes depolarization-evoked exocytosis (measured by membrane capacitance) and hormone secretion (measured biochemically). Single-vesicle optical (by TIRF microscopy) and biophysical measurements of ATP release indicate that G100V/C103V retards initial fusion-pore opening, hinders its expansion and leads to premature closure in most instances. We conclude that the TMD of VAMP2 plays a critical role in membrane fusion and that the structural mobility provided by the central small amino acids is crucial for exocytosis by influencing the molecular re-arrangements of the lipid membrane that are necessary for fusion pore opening and expansion.Benoît HastoyPier A. ScottiAlexandra MilochauZahia Fezoua-BoubegtitenJorge RodasRémi MegretBernard DesbatMichel LaguerreSabine CastanoDavid PerraisPatrik RorsmanReiko OdaJochen LangNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-16 (2017) |
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Medicine R Science Q |
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Medicine R Science Q Benoît Hastoy Pier A. Scotti Alexandra Milochau Zahia Fezoua-Boubegtiten Jorge Rodas Rémi Megret Bernard Desbat Michel Laguerre Sabine Castano David Perrais Patrik Rorsman Reiko Oda Jochen Lang A Central Small Amino Acid in the VAMP2 Transmembrane Domain Regulates the Fusion Pore in Exocytosis |
| description |
Abstract Exocytosis depends on cytosolic domains of SNARE proteins but the function of the transmembrane domains (TMDs) in membrane fusion remains controversial. The TMD of the SNARE protein synaptobrevin2/VAMP2 contains two highly conserved small amino acids, G100 and C103, in its central portion. Substituting G100 and/or C103 with the β-branched amino acid valine impairs the structural flexibility of the TMD in terms of α-helix/β-sheet transitions in model membranes (measured by infrared reflection-absorption or evanescent wave spectroscopy) during increase in protein/lipid ratios, a parameter expected to be altered by recruitment of SNAREs at fusion sites. This structural change is accompanied by reduced membrane fluidity (measured by infrared ellipsometry). The G100V/C103V mutation nearly abolishes depolarization-evoked exocytosis (measured by membrane capacitance) and hormone secretion (measured biochemically). Single-vesicle optical (by TIRF microscopy) and biophysical measurements of ATP release indicate that G100V/C103V retards initial fusion-pore opening, hinders its expansion and leads to premature closure in most instances. We conclude that the TMD of VAMP2 plays a critical role in membrane fusion and that the structural mobility provided by the central small amino acids is crucial for exocytosis by influencing the molecular re-arrangements of the lipid membrane that are necessary for fusion pore opening and expansion. |
| format |
article |
| author |
Benoît Hastoy Pier A. Scotti Alexandra Milochau Zahia Fezoua-Boubegtiten Jorge Rodas Rémi Megret Bernard Desbat Michel Laguerre Sabine Castano David Perrais Patrik Rorsman Reiko Oda Jochen Lang |
| author_facet |
Benoît Hastoy Pier A. Scotti Alexandra Milochau Zahia Fezoua-Boubegtiten Jorge Rodas Rémi Megret Bernard Desbat Michel Laguerre Sabine Castano David Perrais Patrik Rorsman Reiko Oda Jochen Lang |
| author_sort |
Benoît Hastoy |
| title |
A Central Small Amino Acid in the VAMP2 Transmembrane Domain Regulates the Fusion Pore in Exocytosis |
| title_short |
A Central Small Amino Acid in the VAMP2 Transmembrane Domain Regulates the Fusion Pore in Exocytosis |
| title_full |
A Central Small Amino Acid in the VAMP2 Transmembrane Domain Regulates the Fusion Pore in Exocytosis |
| title_fullStr |
A Central Small Amino Acid in the VAMP2 Transmembrane Domain Regulates the Fusion Pore in Exocytosis |
| title_full_unstemmed |
A Central Small Amino Acid in the VAMP2 Transmembrane Domain Regulates the Fusion Pore in Exocytosis |
| title_sort |
central small amino acid in the vamp2 transmembrane domain regulates the fusion pore in exocytosis |
| publisher |
Nature Portfolio |
| publishDate |
2017 |
| url |
https://doaj.org/article/01d582e4c0cb4700b6227323642fc200 |
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