Control of vesicle fusion by a tyrosine phosphatase.
The tyrosine phosphatase PTP-MEG2 is targeted by its amino-terminal Sec14p homology domain to the membrane of secretory vesicles. There it regulates vesicle size by promoting homotypic vesicle fusion by a mechanism that requires its catalytic activity. Here, we identify N-ethylmaleimide-sensitive factor (NSF), a key regulator of vesicle fusion, as a substrate for PTP-MEG2. PTP-MEG2 reduced the phosphotyrosine content of NSF and co-localized with NSF and syntaxin 6 in intact cells. Furthermore, endogenous PTP-MEG2 co-immunoprecipitated with endogenous NSF. Phosphorylation of NSF at Tyr 83, as well as an acidic substitution at the same site, increased its ATPase activity and prevented alphaSNAP binding. Conversely, expression of a Y83F mutant of NSF caused spontaneous fusion events. Our results suggest that the molecular mechanism by which PTP-MEG2 promotes secretory vesicle fusion involves the local release of NSF from a tyrosine-phosphorylated, inactive state. This represents a novel mechanism for localized regulation of NSF and the first demonstrated role for a protein tyrosine phosphatase in the regulated secretory pathway.
Huynh
Huong
H
Program of Inflammation, Infectious and Inflammatory Disease Center, and Program of Signal Transduction, Cancer Center, The Burnham Institute, 10901 North Torrey Pines Road, La Jolla, CA 92037, USA.
Bottini
Nunzio
N
Williams
Scott
S
Cherepanov
Vera
V
Musumeci
Lucia
L
Saito
Kan
K
Bruckner
Shane
S
Vachon
Eric
E
Wang
Xiaodong
X
Kruger
Joshua
J
Chow
Chung-Wai
CW
Pellecchia
Maurizio
M
Monosov
Edvard
E
Greer
Peter A
PA
Trimble
William
W
Downey
Gregory P
GP
Mustelin
Tomas
T
D006801:Humans; D007425:Intracellular Membranes / Q000502:physiology; D019169:Jurkat Cells; D008561:Membrane Fusion; D008565:Membrane Proteins / Q000378:metabolism; D008854:Microscopy, Electron; D052066:N-Ethylmaleimide-Sensitive Proteins; D010766:Phosphorylation; D019000:Phosphotyrosine; D011485:Protein Binding; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism / Q000502:physiology*; D054558:Protein Tyrosine Phosphatases, Non-Receptor; D050765:Qa-SNARE Proteins; D022142:Secretory Vesicles / Q000201:enzymology / Q000502:physiology*; D033921:Vesicular Transport Proteins / Q000378:metabolism
D008565:Membrane Proteins; D050765:Qa-SNARE Proteins; D033921:Vesicular Transport Proteins; D019000:Phosphotyrosine; C447615:PTPN9 protein, human; D017027:Protein Tyrosine Phosphatases; D054558:Protein Tyrosine Phosphatases, Non-Receptor; D052066:N-Ethylmaleimide-Sensitive Proteins