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 · Tue Jan 10 00:00:00 EST 1989 · Biochemistry; (USA)

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

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

---

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), Vol. 28:1; ISSN 0006-2960
• Country of Publication: United States
• Language: English

Related Subjects

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