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ADP ribosylation factor 1 mutants identify a phospholipase D effector region and reveal that phospholipase D participates in lysosomal secretion but is not sufficient for recruitment of coatomer I.
- Source :
-
The Biochemical journal [Biochem J] 1999 Jul 01; Vol. 341 ( Pt 1), pp. 185-92. - Publication Year :
- 1999
-
Abstract
- The small GTP-binding protein, ADP-ribosylation factor 1 (ARF1) is essential for the formation of coatomer-coated vesicles from the Golgi and is also an activator of phospholipase D (PLD). Moreover, ARF1-regulated PLD is part of the signal-transduction pathway that can lead to secretion. In this study, substitution and deletion mutants of ARF1 were tested for their ability to activate PLD. These map the PLD effector region of ARF1 to the alpha2 helix, part of the beta2-strand and the N-terminal helix and its ensuing loop. ARF mutants with an increased or decreased ability to activate PLD showed similar characteristics when tested for their ability to stimulate secretion from HL60 cells. ARF1, deleted of the N-terminal 17 amino acid residues (Ndel17), did not support PLD activity or secretion, and neither did it inhibit the activity of wild-type myristoylated ARF1 (myrARF1). In contrast, Ndel17 effectively competed with wild-type myrARF1 to prevent coatomer binding to membranes. This appears to define a structural role for Ndel17, as it can bind a high-molecular mass complex in cytosol. In addition, ethanol has no effect on recruitment of coatomer to membrane. We conclude that the function of ARF-regulated PLD is in the signal-transduction pathway leading to secretion of lysosomal granules, and not as an essential component of ARF1-mediated coatomer binding.
- Subjects :
- ADP-Ribosylation Factor 1
ADP-Ribosylation Factors
Amino Acid Sequence
Biological Transport
Cell Fractionation
Cell Membrane Permeability
Coated Vesicles metabolism
Coatomer Protein
DNA Mutational Analysis
Enzyme Activation
Ethanol pharmacology
GTP-Binding Proteins chemistry
GTP-Binding Proteins genetics
Golgi Apparatus metabolism
Guanosine 5'-O-(3-Thiotriphosphate) metabolism
HL-60 Cells
Humans
Membrane Proteins metabolism
Models, Molecular
Molecular Sequence Data
Mutation
Myristic Acid metabolism
Protein Binding drug effects
Protein Processing, Post-Translational
Protein Structure, Secondary
Sequence Deletion
Signal Transduction
GTP-Binding Proteins metabolism
Lysosomes metabolism
Phospholipase D metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0264-6021
- Volume :
- 341 ( Pt 1)
- Database :
- MEDLINE
- Journal :
- The Biochemical journal
- Publication Type :
- Academic Journal
- Accession number :
- 10377261