Munc18-1 is critical for plasma membrane localization of syntaxin1 but not of SNAP-25 in PC12 cells.
Although Munc18-1 was originally identified as a syntaxin1-interacting protein, the physiological significance of this interaction remains unclear. In fact, recent studies of Munc18-1 mutants have suggested that Munc18-1 plays a critical role for docking of secretory vesicles, independent of syntaxin1 regulation. Here we investigated the role of Munc18-1 in syntaxin1 localization by generating stable neuroendocrine cell lines in which Munc18-1 was strongly down-regulated. In these cells, the secretion capability, as well as the docking of dense-core vesicles, was significantly reduced. More importantly, not only was the expression level of syntaxin1 reduced, but the localization of syntaxin1 at the plasma membrane was also severely perturbed. The mislocalized syntaxin1 resided primarily in the perinuclear region of the cells, in which it was highly colocalized with Secretogranin II, a marker protein for dense-core vesicles. In contrast, the expression level and the plasma membrane localization of SNAP-25 were not affected. Furthermore, the syntaxin1 localization and the secretion capability were restored upon transfection-mediated reintroduction of Munc18-1. Our results indicate that endogenous Munc18-1 plays a critical role for the plasma membrane localization of syntaxin1 in neuroendocrine cells and therefore necessitates the interpretation of Munc18-1 mutant phenotypes to be in terms of mislocalized syntaxin1.
Arunachalam
Lakshmanan
L
Division of Fundamental Neurobiology, Toronto Western Research Institute, University Health Network, Toronto, Ontario, M5T 2S8, Canada.
Han
Liping
L
Tassew
Nardos G
NG
He
Yu
Y
Wang
Li
L
Xie
Li
L
Fujita
Yoshihito
Y
Kwan
Edwin
E
Davletov
Bazbek
B
Monnier
Philippe P
PP
Gaisano
Herbert Y
HY
Sugita
Shuzo
S
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D018613:Microscopy, Confocal; D051938:Munc18 Proteins / Q000235:genetics / Q000378:metabolism*; D020932:Nerve Growth Factor / Q000494:pharmacology; D016716:PC12 Cells; D021381:Protein Transport / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D022142:Secretory Vesicles / Q000187:drug effects / Q000648:ultrastructure; D013347:Subcellular Fractions / Q000378:metabolism; D050825:Synaptosomal-Associated Protein 25 / Q000378:metabolism*; D050827:Syntaxin 1 / Q000378:metabolism*; D014162:Transfection
D051938:Munc18 Proteins; D012333:RNA, Messenger; C497939:Stxbp1 protein, rat; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1; D020932:Nerve Growth Factor