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Characterization of the regulation of phospholipase D activity in the detergent-insoluble fraction of HL60 cells by protein kinase C and small G-proteins.
- Source :
-
The Biochemical journal [Biochem J] 1999 Apr 01; Vol. 339 ( Pt 1), pp. 87-93. - Publication Year :
- 1999
-
Abstract
- Phospholipase D (PLD) activity has been shown to be GTP-dependent both in vivo and in vitro. One protein that confers GTP sensitivity to PLD activity in vitro is the low-molecular-mass G-protein ADP-ribosylation factor (Arf). However, members of the Rho family and protein kinase C (PKC) have also been reported to activate PLD in various cell systems. We have characterized the stimulation of PLD in HL60 cell membranes by these proteins. The results demonstrate that a considerable proportion of HL60 PLD activity is located in a detergent-insoluble fraction of the cell membrane that is unlikely to be a caveolae-like domain, but is probably cytoskeletal. This PLD activity required the presence of Arf1, a Rho-family member and PKC for efficient catalysis of the lipid substrate, suggesting that the activity represents PLD1. We show that recombinant human PLD1b is regulated in a similar manner to HL60-membrane PLD, and that PKCalpha and PKCdelta are equally effective PLD activators. Therefore maximum PLD activity requires Arf, a Rho-family member and PKC, emphasizing the high degree of regulation of this enzyme.
- Subjects :
- ADP-Ribosylation Factor 1
ADP-Ribosylation Factors
Cell Membrane enzymology
Cloning, Molecular
Detergents
Enzyme Activation
HL-60 Cells
Humans
Phospholipase D genetics
Recombinant Fusion Proteins genetics
Recombinant Fusion Proteins metabolism
Solubility
GTP-Binding Proteins metabolism
Phospholipase D metabolism
Protein Kinase C metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0264-6021
- Volume :
- 339 ( Pt 1)
- Database :
- MEDLINE
- Journal :
- The Biochemical journal
- Publication Type :
- Academic Journal
- Accession number :
- 10085231