Molecular Biology of the Cell click for CBE Life Science Education Page

Home Help [Feedback] [For Subscribers] [Archive] [Search] --
QUICK SEARCH:   [advanced]


     

MBC in Press, published online ahead of print October 7, 2009
Mol. Biol. Cell 10.1091/mbc.E09-08-0712

A more recent version of this article appeared on December 1, 2009
This Article
Right arrow Full Text (PDF)
Right arrow Supplemental Materials
Right arrow All Versions of this Article:
E09-08-0712v1
20/23/4962    most recent
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Google Scholar
Right arrow Articles by Han, L.
Right arrow Articles by Sugita, S.
PubMed
Right arrow PubMed Citation
Right arrow Articles by Han, L.
Right arrow Articles by Sugita, S.

Submitted on August 19, 2009
Revised on September 22, 2009
Accepted on September 25, 2009

Rescue of Munc18–1 and -2 Double Knockdown Reveals the Essential Functions of Interaction between Munc18 and Closed Syntaxin in PC12 Cells

Liping Han,*{dagger} Tiandan Jiang,*{dagger} Gayoung A. Han,*{dagger} Nancy T. Malintan,{ddagger} Li Xie,{dagger}{sect} Li Wang,* Frederick W. Tse,|| Herbert Y. Gaisano,*{dagger}{sect} Brett M. Collins,¶ Frederic A. Meunier,{ddagger} and Shuzo Sugita*{dagger}

*Division of Fundamental Neurobiology, University Health Network, Toronto, Ontario, M5T 2S8, Canada; Departments of {dagger}Physiology and {sect}Medicine, Faculty of Medicine, University of Toronto, Toronto, Ontario, M5S 1A8, Canada; {ddagger}Queensland Brain Institute and School of Biomedical Science and Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia 4072; ||Department of Pharmacology, University of Alberta, Edmonton, Alberta, Canada T6G 2H7

Monitoring Editor: Keith E. Mostov

Munc18–1 binds to syntaxin-1A via two distinct sites referred to as the ‘closed’ conformation and N-terminus binding. The latter has been shown to stimulate SNARE-mediated exocytosis, whereas the former is believed to be inhibitory or dispensable. To precisely define the contributions of each binding mode, we have engineered Munc18–1/-2 double knockdown neurosecretory cells and show that not only syntaxin-1A and -1B but also syntaxin-2 and -3 were significantly reduced as a result of Munc18–1 and -2 knockdown. Syntaxin-1 was mislocalized and the regulated secretion was abolished. We next examined the abilities of Munc18–1 mutants to rescue the defective phenotypes. Mutation (K46E/E59K) of Munc18–1 that selectively prevents binding to ‘closed’ syntaxin-1 was unable to restore syntaxin-1 expression, localization or secretion. In contrast, mutations (F115E/E132A) of Munc18–1 that selectively impair binding to the syntaxin-1 N-terminus could still rescue the defective phenotypes. Our results indicate that Munc18–1 and -2 act in concert to support the expression of a broad range of syntaxins and to deliver syntaxin-1 to the plasma membrane. Our studies also indicate that the binding to the ‘closed’ conformation of syntaxin is essential for Munc18–1 stimulatory action while the binding to syntaxin N-terminus plays a more limited role in neurosecretory cells.

Address correspondence to: Shuzo Sugita (ssugita{at}uhnres.utoronto.ca)






Home Help [Feedback] [For Subscribers] [Archive] [Search] --
Copyright © 2009 by The American Society for Cell Biology. Terms of copyright protection, warranties, and disclaimers.