Inhibition of Phospholipase D by Amphiphysins

Two distinct proteins inhibiting phospholipase D (PLD) activity in rat brain cytosol were previously purified and identified as synaptojanin and AP180, which are specific to nerve terminals and associate with the clathrin coat. Two additional PLD-inhibitory proteins have now been purified and identi...

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Bibliographic Details
Published in:The Journal of biological chemistry 2000-06, Vol.275 (25), p.18751-18758
Main Authors: Lee, Chunghee, Kim, Seung Ryul, Chung, Joon-Ki, Frohman, Michael A., Kilimann, Manfred W., Rhee, Sue Goo
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
Cell Line
Chromatography, DEAE-Cellulose
Chromatography, High Pressure Liquid
Clathrin - metabolism
COS Cells
Cricetinae
Endocytosis
Enzyme Inhibitors - pharmacology
Glutathione Transferase - chemistry
Isoenzymes - antagonists & inhibitors
Molecular Sequence Data
Nerve Tissue Proteins - chemistry
Nerve Tissue Proteins - isolation & purification
Nerve Tissue Proteins - pharmacology
Phospholipase D - antagonists & inhibitors
Rats
Recombinant Fusion Proteins - chemistry
Recombinant Fusion Proteins - pharmacology
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Summary:Two distinct proteins inhibiting phospholipase D (PLD) activity in rat brain cytosol were previously purified and identified as synaptojanin and AP180, which are specific to nerve terminals and associate with the clathrin coat. Two additional PLD-inhibitory proteins have now been purified and identified as the amphiphysins I and II, which forms a heterodimer that also associates with the clathrin coat. Bacterially expressed recombinant amphiphysins inhibited both PLD1 and PLD2 isozymes in vitro with a potency similar to that of brain amphiphysin (median inhibitory concentration of ∼15 nm). Expressions of either amphiphysin in COS-7 cells reduced activity of endogenous PLD as well as exogenously expressed PLD1 and PLD2. Coprecipitation experiments suggested that the inhibitory effect of amphiphysins results from their direct interaction with PLDs. The NH2 terminus of amphiphysin I was critical for both inhibition of and binding to PLD. Phosphatidic acid formed by signal-induced PLD is thought to be required for the assembly of clathrin-coated vesicles during endocytosis. Thus, the inhibition of PLD by amphiphysins, synaptojanin, and AP180 might play an important role in synaptic vesicle trafficking.
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.M001695200