Plasma membrane associated phospholipase C from human platelets: Synergistic stimulation of phosphatidylinositol 4,5-bisphosphate hydrolysis by thrombin and guanosine 5 prime -O-(3-thiotriphosphate)

Journal Article · Mon Jan 09 23:00:00 EST 1989 · Biochemistry; (USA)

DOI:https://doi.org/10.1021/bi00427a009· OSTI ID:5413331

Baldassare, J J; Henderson, P A; Fisher, G J ^[1]

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1. American Red Cross, St. Louis, MO (USA)

The effects of thrombin and GTP{gamma}S on the hydrolysis of phosphoinositides by membrane-associated phospholipase C (PLC) from human platelets were examined with endogenous ({sup 3}H)inositol-labeled membranes or with lipid vesicles containing either ({sup 3}H)phosphatidylinositol or ({sup 3}H)phosphatidylinositol 4,5-bisphosphate. GTP{gamma}S (1 {mu}M) or thrombin (1 unit/mL) did not stimulate release of inositol trisphosphate (IP{sub 3}), inositol bisphosphate (IP{sub 2}), or inositol phosphate (IP) from ({sup 3}H)inositol-labeled membranes. IP{sub 2} and IP{sub 3}, but not IP, from ({sup 3}H)inositol-labeled membranes were, however, stimulated 3-fold by GTP{gamma}S (1 {mu}M) plus thrombin (1 unit/mL). A higher concentration of GTP{gamma}S (100 {mu}M) alone also stimulated IP{sub 2} and IP{sub 3}, but not IP, release. In the presence of 1 mM calcium, release of IP{sub 2} and IP{sub 3} was increased 6-fold over basal levels; however, formation of IP was not observed. At submicromolar calcium concentration, hydrolysis of exogenous phosphatidylinositol 4,5-bisphosphate (PIP{sub 2}) by platelet membrane associated PLC was also markedly enhanced by GTP{gamma}S (100 {mu}M) or GTP{gamma}S (1 {mu}M) plus thrombin (1 unit/mL). Under identical conditions, exogenous phosphatidylinositol (PI) was not hydrolyzed. The same substrate specificity was observed when the membrane-associated PLC was activated with 1 mM calcium. Thrombin-induced hydrolysis of PIP{sub 2} was inhibited by treatment of the membranes with pertussis toxin or pretreatment of intact platelets with 12-O-tetradecanoyl-13-acetate (TPA) prior to preparation of membranes. Pertussis toxin did not inhibit GTP{gamma}S (100 {mu}M) or calcium (1 mM) dependent PIP{sub 2} breakdown, while TPA inhibited GTP{gamma}S-dependent but not calcium-dependent phospholipase C activity.

• OSTI ID: 5413331
• Journal Information: Biochemistry; (USA), Journal Name: Biochemistry; (USA) Vol. 28:1; ISSN 0006-2960; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ALKALINE EARTH METAL COMPOUNDS
ANIMALS
AROMATICS
AZAARENES
BIOCHEMISTRY
BIOLOGICAL EFFECTS
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BLOOD PLATELETS
BODY FLUIDS
CALCIUM COMPOUNDS
CELL CONSTITUENTS
CELL MEMBRANES
CHEMICAL REACTIONS
CHEMISTRY
COAGULANTS
DECOMPOSITION
DRUGS
ENZYMES
ESTERS
GUANOSINE
HEMATOLOGIC AGENTS
HEMOSTATICS
HETEROCYCLIC COMPOUNDS
HYDROGEN COMPOUNDS
HYDROLASES
HYDROLYSIS
LIPASE
LIPIDS
LYSIS
MAMMALS
MAN
MATERIALS
MEMBRANES
NUCLEOSIDES
NUCLEOTIDES
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
PEPTIDE HYDROLASES
PH VALUE
PHOSPHOLIPIDS
PRIMATES
PURINES
RIBOSIDES
SERINE PROTEINASES
SOLVOLYSIS
THROMBIN
TRITIUM COMPOUNDS
VERTEBRATES