GENERIC 11018470

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Predicate	Object
rdf:type	pubmed:Citation
lifeskim:mentions	umls-concept:C0031670, umls-concept:C0033634, umls-concept:C0079469, umls-concept:C0086418, umls-concept:C0815000, umls-concept:C1416956, umls-concept:C1879547
pubmed:issue	2-3
pubmed:dateCreated	2000-11-28
pubmed:abstractText	Regulation of phospholipase D (PLD) activity participating in signal transduction involves complex interactions with small G-proteins (ARF, Rho) and protein kinase C isoforms (PKCalpha). In SK-N-MC human neuroblastoma cells, phorbol ester (TPA) activation of PLD was enhanced by overexpressing myristoylated alanine-rich C kinase substrate (MARCKS). To study MARCKS interactions with PLD, we investigated PLD isoform expression and activation by TPA and GTPgammaS in intact and digitonin-permeabilized clones transfected with MARCKS (M22). PLD2 was in both cytosol and membrane fractions while PLD1 was primarily membrane-associated in both vector control and M22 cells; location or quantities were unaltered by TPA treatment. TPA-stimulated PLD activity was higher in both intact and digitonin-permeabilized M22 cells than in vector controls. In contrast, GTPgammaS-stimulated PLD activity was independent of MARCKS expression but was additive with MARCKS-PKC-dependent activation in permeabilized cells. Combinations of PKC inhibition and down-regulation in intact and permeabilized (with GTPgammaS present) cells indicated that a PKC-mediated phosphorylation event was necessary in intact cells without access to GTPgammaS, stimulation of PLD mediated by GTPgammaS was independent of PKC, and PLD activation by PKC in permeabilized cells was kinase-independent. Western blot analysis showed that MARCKS, PKCalpha, PLD1 and PLD2 were present in a detergent-insoluble fraction (DIF); GTPgammaS increased recovery of PLD2 in DIF. Disruption of cholesterol-rich DIFs with digitonin, cyclodextrin or filipin potentiated activation of PLD by TPA. Our studies suggest that activation of PLD by PKC requires MARCKS and can involve both phosphorylation-independent and -dependent processes. As PLD activation by GTPgammaS is PKC-MARCKS-independent, MARCKS may provide a fine tuning component in conjunction with G-protein-mediated mechanisms for regulation of PLD.
pubmed:language	eng
pubmed:journal	http://linkedlifedata.com/resource/pubmed/journal/0217513
pubmed:citationSubset	IM
pubmed:chemical	http://linkedlifedata.com/resource/pubmed/chemical/Cyclodextrins, http://linkedlifedata.com/resource/pubmed/chemical/Digitonin, http://linkedlifedata.com/resource/pubmed/chemical/Filipin, http://linkedlifedata.com/resource/pubmed/chemical/Guanosine 5'-O-(3-Thiotriphosphate), http://linkedlifedata.com/resource/pubmed/chemical/Intracellular Signaling Peptides..., http://linkedlifedata.com/resource/pubmed/chemical/Membrane Proteins, http://linkedlifedata.com/resource/pubmed/chemical/Phospholipase D, http://linkedlifedata.com/resource/pubmed/chemical/Protein Kinase C, http://linkedlifedata.com/resource/pubmed/chemical/Tetradecanoylphorbol Acetate, http://linkedlifedata.com/resource/pubmed/chemical/beta-Cyclodextrins, http://linkedlifedata.com/resource/pubmed/chemical/betadex, http://linkedlifedata.com/resource/pubmed/chemical/myristoylated alanine-rich C..., http://linkedlifedata.com/resource/pubmed/chemical/phospholipase D1, http://linkedlifedata.com/resource/pubmed/chemical/phospholipase D2
pubmed:status	MEDLINE
pubmed:month	Sep
pubmed:issn	0006-3002
pubmed:author	pubmed-author:ByersD MDM, pubmed-author:CookH WHW, pubmed-author:MorashS CSC
pubmed:issnType	Print
pubmed:day	27
pubmed:volume	1487
pubmed:owner	NLM
pubmed:authorsComplete	Y
pubmed:pagination	177-89
pubmed:dateRevised	2007-11-15
pubmed:meshHeading	pubmed-meshheading:11018470-Animals, pubmed-meshheading:11018470-Cell Fractionation, pubmed-meshheading:11018470-Cyclodextrins, pubmed-meshheading:11018470-Digitonin, pubmed-meshheading:11018470-Electroporation, pubmed-meshheading:11018470-Enzyme Activation, pubmed-meshheading:11018470-Filipin, pubmed-meshheading:11018470-Guanosine 5'-O-(3-Thiotriphosphate), pubmed-meshheading:11018470-Humans, pubmed-meshheading:11018470-Intracellular Signaling Peptides and Proteins, pubmed-meshheading:11018470-Membrane Proteins, pubmed-meshheading:11018470-Phospholipase D, pubmed-meshheading:11018470-Protein Biosynthesis, pubmed-meshheading:11018470-Protein Kinase C, pubmed-meshheading:11018470-Rats, pubmed-meshheading:11018470-Signal Transduction, pubmed-meshheading:11018470-Tetradecanoylphorbol Acetate, pubmed-meshheading:11018470-Tumor Cells, Cultured, pubmed-meshheading:11018470-beta-Cyclodextrins
pubmed:year	2000
pubmed:articleTitle	Activation of phospholipase D by PKC and GTPgammaS in human neuroblastoma cells overexpressing MARCKS.
pubmed:affiliation	Atlantic Research Centre, Departments of Pediatrics and Biochemistry and Molecular Biology, Dalhousie University, Halifax, NS, Canada.
pubmed:publicationType	Journal Article, Research Support, Non-U.S. Gov't