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PMID: 2173906 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Guanosine 5'-[gamma-thio]triphosphate-stimulated hydrolysis of phosphatidylinositol 4,5-bisphosphate in HL-60 granulocytes. Evidence that the guanine nucleotide acts by relieving phospholipase C from an inhibitory constraint.

The Biochemical journal ·Vol. 271 ·No. 3 ·1990-11-01 ·Pages 743-8

Camps M, Hou CF, Jakobs KH, Gierschik P

Abstract

Myeloid differentiated human leukaemia (HL-60) cells contain a soluble phospholipase C that hydrolysed phosphatidylinositol 4.5-bisphosphate and was markedly stimulated by the metabolically stable GTP analogue guanosine 5'-[gamma-thio]triphosphate (GTP[S]). Half-maximal and maximal (up to 5-fold) stimulation of inositol phosphate formation by GTP[S] occurred at 1.5 microM and 30 microM respectively. Other nucleotides (GTP, GDP, GMP, guanosine 5'-[beta-thio]diphosphate. ATP, adenosine 5'-[gamma-thio]triphosphate, UTP) did not affect phospholipase C activity, GTP[S] stimulation of inositol phosphate accumulation was inhibited by excess GDP, but not by ADP. The effect of GTP[S] on inositol phosphate formation was absolutely dependent on and markedly stimulated by free Ca2+ (median effective concn. approximately 100 nM). Analysis of inositol phosphates by anion-exchange chromatography revealed InsP3 as the major product of GTP[S]-stimulated phospholipase C activity. In the absence of GTP[S], specific phospholipase C activity was markedly decreased when tested at high protein concentrations, whereas GTP[S] stimulation of the enzyme was markedly enhanced under these conditions. As both basal and GTP[S]-stimulated inositol phosphate formation were linear with time whether studied at low or high protein concentration, these results suggest that (a) phospholipase C is under an inhibitory constraint and (b) GTP[S] relieves this inhibition, most likely by activating a soluble GTP-binding protein.

MeSH Terms
Cytosol/enzymology,metabolism Deoxycholic Acid/pharmacology Enzyme Activation Granulocytes/enzymology,metabolism Guanosine 5'-O-(3-Thiotriphosphate)/pharmacology Guanosine Diphosphate/pharmacology Humans Hydrogen-Ion Concentration Hydrolysis/drug effects Inositol Phosphates/biosynthesis,blood Kinetics Leukemia, Experimental/blood,pathology Leukemia, Myeloid/blood,pathology Phosphatidylinositol 4,5-Diphosphate Phosphatidylinositols/biosynthesis,blood Phospholipids/blood Stimulation, Chemical Tumor Cells, Cultured Type C Phospholipases/antagonists & inhibitors,blood
Chemicals
Inositol Phosphates Phosphatidylinositol 4,5-Diphosphate Phosphatidylinositols Phospholipids Deoxycholic Acid Guanosine Diphosphate Guanosine 5'-O-(3-Thiotriphosphate) Type C Phospholipases
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Camps M
Pharmakologisches Institut, Universität Heidelberg, Federal Republic of Germany.
Hou C F
Jakobs K H
Gierschik P
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Article Info
Journal
The Biochemical journal
Abbr.
Biochem J
ISSN
0264-6021
DOI
10.1042/bj2710743
Published
1990-11-01
Pages
743-8
Language
English
Region
England
NLM ID
2984726R
PMCID
PMC1149625
Subset
IM
External Links
PubMed source DOI: 10.1042/bj2710743 PMC Full Text View journal details
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