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PMID: 1700902 Published · ppublish English Comparative Study Journal Article Research Support, Non-U.S. Gov't

Functional relationship between cyclic AMP-dependent protein phosphorylation and platelet inhibition.

The Biochemical journal ·Vol. 271 ·No. 3 ·1990-11-01 ·Pages 815-9

Siess W, Lapetina EG

Abstract

Exposure of human platelets to prostacyclin (PGI2), iloprost or prostaglandin E1 (PGE1) elicits the cyclic AMP-dependent phosphorylation of proteins of 22, 24, 30, 39, 50, 60 and 250 kDa (P22, P24 etc.). P22 was recently identified as rap 1B, a ras-like protein, and P24 was shown to be the beta-chain of glycoprotein Ib. We found that cyclic AMP-dependent phosphorylation of all proteins except P22 was maximal 1 min after exposure of platelets to PGI2, iloprost or PGE1; maximal phosphorylation of P22 occurred after 45 min of incubation. Inhibition of thrombin-induced platelet activation required only a 30 s incubation with PGI2 or iloprost; at this time phosphorylation of P22 was only slightly increased. Although at maximal concentrations PGI2 was more potent than PGE1 in inhibiting thrombin-induced platelet activation, no difference in the degree and the kinetics of cyclic AMP-dependent protein phosphorylation was found. Platelets that had been preincubated and washed in the presence of PGE1 and later resuspended in the absence of PGE1 responded fully to activation by thrombin despite maximal phosphorylation of P22 and P24. Furthermore, addition of PGI2 to PGE1-washed platelets prevented thrombin-induced platelet activation, but did not evoke further phosphorylation of P22 or P24. Phosphorylation of P39 and P50 correlated better with PGI2-induced inhibition of platelet activation. In experiments in which PGE1-induced inhibition of platelet activation was overcome by the addition of thrombin, no dephosphorylation of proteins phosphorylated by cyclic AMP-dependent kinases was observed. These experiments indicate that: (a) phosphorylation of rap 1B and glycoprotein Ib is not related to platelet inhibition by cyclic AMP; (b) phosphorylation of other proteins such as P39 and P50 probably plays a role in mediating cyclic AMP-dependent platelet inhibition; (c) reactions other than cyclic AMP-dependent protein phosphorylation may participate in platelet inhibition by cyclic AMP.

MeSH Terms
Alprostadil/pharmacology Blood Proteins/metabolism Epoprostenol/pharmacology Humans Iloprost/pharmacology Molecular Weight Platelet Activation/drug effects,physiology Platelet Aggregation Inhibitors/pharmacology Protein Kinases/blood,pharmacology,physiology
Chemicals
Blood Proteins Platelet Aggregation Inhibitors Epoprostenol Protein Kinases Alprostadil Iloprost
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Siess W
Division of Cell Biology, Burroughs Wellcome Co., Research Triangle Park, NC 27709.
Lapetina E G
References (35)
35 references, click to expand
  1. A ras-related protein is phosphorylated and translocated by agonists that increase cAMP levels in human platelets.
    Proc Natl Acad Sci U S A. 1989 May;86(9):3131-4 PMID: 2470091
  2. Prostacyclin inhibits platelet aggregation induced by phorbol ester or Ca2+ ionophore at steps distal to activation of protein kinase C and Ca2+-dependent protein kinases.
    Biochem J. 1989 Feb 15;258(1):57-65 PMID: 2494992
  3. Phosphorylation of smg p21, a ras p21-like GTP-binding protein, by cyclic AMP-dependent protein kinase in a cell-free system and in response to prostaglandin E1 in intact human platelets.
    J Biol Chem. 1989 Sep 15;264(26):15688-95 PMID: 2504724
  4. Molecular mechanisms of platelet activation.
    Physiol Rev. 1989 Jan;69(1):58-178 PMID: 2536184
  5. Phosphorylation by cyclic AMP-dependent protein kinase of a human platelet Mr 22,000 GTP-binding protein (smg p21) having the same putative effector domain as the ras gene products.
    Biochem Biophys Res Commun. 1988 Dec 30;157(3):851-60 PMID: 2849942
  6. Prostacyclin analogues reduce ADP-ribosylation of the alpha-subunit of the regulatory Gs-protein and diminish adenosine (A2) responsiveness of platelets.
    Br J Pharmacol. 1987 Mar;90(3):501-10 PMID: 2436701
  7. Platelet cytosolic 44-kDa protein is a substrate of cholera toxin-induced ADP-ribosylation and is not recognized by antisera against the alpha subunit of the stimulatory guanine nucleotide-binding regulatory protein.
    Proc Natl Acad Sci U S A. 1988 Aug;85(16):5899-902 PMID: 3137562
  8. Human cDNAs rap1 and rap2 homologous to the Drosophila gene Dras3 encode proteins closely related to ras in the 'effector' region.
    Oncogene. 1988 Aug;3(2):201-4 PMID: 3045729
  9. Phorbol ester stimulates calcium sequestration in saponized human platelets.
    J Biol Chem. 1987 Nov 25;262(33):16048-54 PMID: 3119582
  10. cAMP-dependent protein kinase. Model for an enzyme family.
    J Biol Chem. 1989 May 25;264(15):8443-6 PMID: 2656679
  11. Solubilization of prostacyclin membrane receptors from human platelets.
    J Biol Chem. 1989 Jan 5;264(1):61-7 PMID: 2642482
  12. Purification of a vasodilator-regulated phosphoprotein from human platelets.
    Eur J Biochem. 1989 Oct 20;185(1):41-50 PMID: 2806262
  13. Release of Ca2+ by inositol 1,4,5-trisphosphate in platelet membrane vesicles is not dependent on cyclic AMP-dependent protein kinase.
    Biochem J. 1989 Feb 1;257(3):715-21 PMID: 2784669
  14. Rap1-B is phosphorylated by protein kinase A in intact human platelets.
    Biochem Biophys Res Commun. 1990 Jul 31;170(2):944-50 PMID: 1696481
  15. Thrombolamban, the 22-kDa platelet substrate of cyclic AMP-dependent protein kinase, is immunologically homologous with the Ras family of GTP-binding proteins.
    Proc Natl Acad Sci U S A. 1990 Jan;87(2):758-62 PMID: 1689052
  16. Stoichiometric and reversible phosphorylation of a 46-kDa protein in human platelets in response to cGMP- and cAMP-elevating vasodilators.
    J Biol Chem. 1990 Feb 25;265(6):3088-93 PMID: 2154470
  17. Structure of catabolite gene activator protein at 2.9 A resolution suggests binding to left-handed B-DNA.
    Nature. 1981 Apr 30;290(5809):744-9 PMID: 6261152
  18. Cyclic nucleotides control a system which regulates Ca2+ sensitivity of platelet secretion.
    Nature. 1984 May 3-9;309(5963):66-8 PMID: 6201746
  19. Regulation of the intracellular calcium level in human blood platelets: cyclic adenosine 3',5'-monophosphate dependent phosphorylation of a 22,000 dalton component in isolated Ca2+-accumulating vesicles.
    Biochim Biophys Acta. 1979 Dec 12;558(3):344-7 PMID: 228723
  20. Effects of collagen, ionophore A23187 and prostaglandin E1 on the phosphorylation of specific proteins in blood platelets.
    Biochem J. 1979 Feb 15;178(2):397-406 PMID: 375934
  21. An enzyme isolated from arteries transforms prostaglandin endoperoxides to an unstable substance that inhibits platelet aggregation.
    Nature. 1976 Oct 21;263(5579):663-5 PMID: 802670
  22. A ras-related gene with transformation suppressor activity.
    Cell. 1989 Jan 13;56(1):77-84 PMID: 2642744
  23. Cyclic AMP turnover in response to prostaglandins in intact platelets: evidence for separate stimulatory and inhibitory prostaglandin receptors.
    Second Messengers Phosphoproteins. 1988;12(1):45-57 PMID: 2848122
  24. Demonstration of cGMP-dependent protein kinase and cGMP-dependent phosphorylation in cell-free extracts of platelets.
    Eur J Biochem. 1986 Jul 1;158(1):203-10 PMID: 3015608
  25. Inhibition of platelet-activating-factor-induced human platelet activation by prostaglandin D2. Differential sensitivity of platelet transduction processes and functional responses to inhibition by cyclic AMP.
    Biochem J. 1985 Nov 15;232(1):267-71 PMID: 3002327
  26. Cyclic AMP and other signals controlling cell development and differentiation in Dictyostelium.
    Annu Rev Biochem. 1987;56:853-79 PMID: 3039908
  27. cAMP reduces the affinity of Ca2+-triggered secretion in platelets.
    FEBS Lett. 1987 May 4;215(1):183-6 PMID: 3032688
  28. Cyclic AMP inhibits platelet activation independently of its effect on cytosolic free calcium.
    Biochem Biophys Res Commun. 1985 Feb 28;127(1):339-45 PMID: 2983714
  29. Evidence against direct involvement of cyclic AMP-dependent protein phosphorylation in the exocytosis of amylase.
    Biochem J. 1988 Dec 15;256(3):867-71 PMID: 2464999
  30. Comparison of equimolar concentrations of iloprost, prostacyclin, and prostaglandin E1 on human platelet function.
    J Lab Clin Med. 1987 Feb;109(2):184-90 PMID: 2433373
  31. Inhibition by forskolin of cytosolic calcium rise, shape change and aggregation in quin2-loaded human platelets.
    FEBS Lett. 1985 Aug 19;188(1):135-40 PMID: 2410292
  32. In situ phosphorylation of platelet actin-binding protein by cAMP-dependent protein kinase stabilizes it against proteolysis by calpain.
    J Biol Chem. 1989 Aug 25;264(24):14282-9 PMID: 2547793
  33. Cyclic AMP-dependent phosphorylation of glycoprotein Ib inhibits collagen-induced polymerization of actin in platelets.
    J Biol Chem. 1989 Jun 5;264(16):9520-6 PMID: 2542312
  34. Platelet glycoprotein Ib beta is phosphorylated on serine 166 by cyclic AMP-dependent protein kinase.
    J Biol Chem. 1989 Sep 15;264(26):15656-61 PMID: 2504723
  35. Protein modification: new clue to Ras lipid glue.
    Nature. 1989 Oct 5;341(6241):384-5 PMID: 2677741
Article Info
Journal
The Biochemical journal
Abbr.
Biochem J
ISSN
0264-6021
DOI
10.1042/bj2710815
Published
1990-11-01
Pages
815-9
Language
English
Region
England
NLM ID
2984726R
PMCID
PMC1149637
Subset
IM
External Links
PubMed source DOI: 10.1042/bj2710815 PMC Full Text View journal details
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