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The nature of the Syntaxin4 C-terminus affects Munc18c-supported SNARE assembly

Vesicular transport of cellular cargo requires targeted membrane fusion and formation of a SNARE protein complex that draws the two apposing fusing membranes together. Insulin-regulated delivery and fusion of glucose transporter-4 storage vesicles at the cell surface is dependent on two key proteins...

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Autores principales: Rehman, Asma, Hu, Shu-Hong, Tnimov, Zakir, Whitten, Andrew E., King, Gordon J., Jarrott, Russell J., Norwood, Suzanne J., Alexandrov, Kirill, Collins, Brett M., Christie, Michelle P., Martin, Jennifer L.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5571939/
https://www.ncbi.nlm.nih.gov/pubmed/28841669
http://dx.doi.org/10.1371/journal.pone.0183366
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author Rehman, Asma
Hu, Shu-Hong
Tnimov, Zakir
Whitten, Andrew E.
King, Gordon J.
Jarrott, Russell J.
Norwood, Suzanne J.
Alexandrov, Kirill
Collins, Brett M.
Christie, Michelle P.
Martin, Jennifer L.
author_facet Rehman, Asma
Hu, Shu-Hong
Tnimov, Zakir
Whitten, Andrew E.
King, Gordon J.
Jarrott, Russell J.
Norwood, Suzanne J.
Alexandrov, Kirill
Collins, Brett M.
Christie, Michelle P.
Martin, Jennifer L.
author_sort Rehman, Asma
collection PubMed
description Vesicular transport of cellular cargo requires targeted membrane fusion and formation of a SNARE protein complex that draws the two apposing fusing membranes together. Insulin-regulated delivery and fusion of glucose transporter-4 storage vesicles at the cell surface is dependent on two key proteins: the SNARE integral membrane protein Syntaxin4 (Sx4) and the soluble regulatory protein Munc18c. Many reported in vitro studies of Munc18c:Sx4 interactions and of SNARE complex formation have used soluble Sx4 constructs lacking the native transmembrane domain. As a consequence, the importance of the Sx4 C-terminal anchor remains poorly understood. Here we show that soluble C-terminally truncated Sx4 dissociates more rapidly from Munc18c than Sx4 where the C-terminal transmembrane domain is replaced with a T4-lysozyme fusion. We also show that Munc18c appears to inhibit SNARE complex formation when soluble C-terminally truncated Sx4 is used but does not inhibit SNARE complex formation when Sx4 is C-terminally anchored (by a C-terminal His-tag bound to resin, by a C-terminal T4L fusion or by the native C-terminal transmembrane domain in detergent micelles). We conclude that the C-terminus of Sx4 is critical for its interaction with Munc18c, and that the reported inhibitory role of Munc18c may be an artifact of experimental design. These results support the notion that a primary role of Munc18c is to support SNARE complex formation and membrane fusion.
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spelling pubmed-55719392017-09-09 The nature of the Syntaxin4 C-terminus affects Munc18c-supported SNARE assembly Rehman, Asma Hu, Shu-Hong Tnimov, Zakir Whitten, Andrew E. King, Gordon J. Jarrott, Russell J. Norwood, Suzanne J. Alexandrov, Kirill Collins, Brett M. Christie, Michelle P. Martin, Jennifer L. PLoS One Research Article Vesicular transport of cellular cargo requires targeted membrane fusion and formation of a SNARE protein complex that draws the two apposing fusing membranes together. Insulin-regulated delivery and fusion of glucose transporter-4 storage vesicles at the cell surface is dependent on two key proteins: the SNARE integral membrane protein Syntaxin4 (Sx4) and the soluble regulatory protein Munc18c. Many reported in vitro studies of Munc18c:Sx4 interactions and of SNARE complex formation have used soluble Sx4 constructs lacking the native transmembrane domain. As a consequence, the importance of the Sx4 C-terminal anchor remains poorly understood. Here we show that soluble C-terminally truncated Sx4 dissociates more rapidly from Munc18c than Sx4 where the C-terminal transmembrane domain is replaced with a T4-lysozyme fusion. We also show that Munc18c appears to inhibit SNARE complex formation when soluble C-terminally truncated Sx4 is used but does not inhibit SNARE complex formation when Sx4 is C-terminally anchored (by a C-terminal His-tag bound to resin, by a C-terminal T4L fusion or by the native C-terminal transmembrane domain in detergent micelles). We conclude that the C-terminus of Sx4 is critical for its interaction with Munc18c, and that the reported inhibitory role of Munc18c may be an artifact of experimental design. These results support the notion that a primary role of Munc18c is to support SNARE complex formation and membrane fusion. Public Library of Science 2017-08-25 /pmc/articles/PMC5571939/ /pubmed/28841669 http://dx.doi.org/10.1371/journal.pone.0183366 Text en © 2017 Rehman et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Rehman, Asma
Hu, Shu-Hong
Tnimov, Zakir
Whitten, Andrew E.
King, Gordon J.
Jarrott, Russell J.
Norwood, Suzanne J.
Alexandrov, Kirill
Collins, Brett M.
Christie, Michelle P.
Martin, Jennifer L.
The nature of the Syntaxin4 C-terminus affects Munc18c-supported SNARE assembly
title The nature of the Syntaxin4 C-terminus affects Munc18c-supported SNARE assembly
title_full The nature of the Syntaxin4 C-terminus affects Munc18c-supported SNARE assembly
title_fullStr The nature of the Syntaxin4 C-terminus affects Munc18c-supported SNARE assembly
title_full_unstemmed The nature of the Syntaxin4 C-terminus affects Munc18c-supported SNARE assembly
title_short The nature of the Syntaxin4 C-terminus affects Munc18c-supported SNARE assembly
title_sort nature of the syntaxin4 c-terminus affects munc18c-supported snare assembly
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5571939/
https://www.ncbi.nlm.nih.gov/pubmed/28841669
http://dx.doi.org/10.1371/journal.pone.0183366
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