The Slp4-a linker domain controls exocytosis through interaction with Munc18-1.syntaxin-1a complex.
Synaptotagmin-like protein 4-a (Slp4-a)/granuphilin-a is specifically localized on dense-core vesicles in certain neuroendocrine cells and negatively controls dense-core vesicle exocytosis through specific interaction with Rab27A. However, the precise molecular mechanism of its inhibitory effect on exocytosis has never been elucidated and is still a matter of controversy. Here we show by deletion and chimeric analyses that the linker domain of Slp4-a interacts with the Munc18-1.syntaxin-1a complex by directly binding to Munc18-1 and that this interaction promotes docking of dense-core vesicles to the plasma membrane in PC12 cells. Despite increasing the number of plasma membrane docked vesicles, expression of Slp4-a strongly inhibited high-KCl-induced dense-core vesicle exocytosis. The inhibitory effect by Slp4-a is absolutely dependent on the linker domain of Slp4-a, because substitution of the linker domain of Slp4-a by that of Slp5 (the closest isoform of Slp4-a that cannot bind the Munc18-1.syntaxin-1a complex) completely abrogated the inhibitory effect. Our findings reveal a novel docking machinery for dense-core vesicle exocytosis: Slp4-a simultaneously interacts with Rab27A and Munc18-1 on the dense-core vesicle and with syntaxin-1a in the plasma membrane.
Fukuda Initiative Research Unit, Riken (The Institute of Physical and Chemical Research), Wako, Saitama 351-0198, Japan.
Takashi
T
Tsuboi
Mitsunori
M
Fukuda
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D005089:Exocytosis; D006801:Humans; D008561:Membrane Fusion; D008565:Membrane Proteins / Q000378:metabolism; D051938:Munc18 Proteins / Q000378:metabolism*; D016716:PC12 Cells; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering / Q000235:genetics / Q000494:pharmacology; D051381:Rats; D050827:Syntaxin 1 / Q000378:metabolism*; D022161:Transport Vesicles / Q000378:metabolism*; D033921:Vesicular Transport Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*
D002352:Carrier Proteins; D008565:Membrane Proteins; D051938:Munc18 Proteins; D034741:RNA, Small Interfering; C512779:SYTL4 protein, human; C457772:SYTL5 protein, human; D050827:Syntaxin 1; C512778:Sytl4 protein, rat; D033921:Vesicular Transport Proteins