Thromb Haemost 2021; 121(12): 1628-1636
DOI: 10.1055/a-1481-2663
Cellular Haemostasis and Platelets

Role of Membrane Lipid Rafts in MRP4 (ABCC4) Dependent Regulation of the cAMP Pathway in Blood Platelets

Tiphaine Belleville-Rolland*
1   Service d'hématologie biologique, AH-HP, Hopital Européen Georges Pompidou, Paris, France
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
,
Alexandre Leuci*
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
,
Alexandre Mansour
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
,
Benoit Decouture
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
,
Fanny Martin
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
,
Sonia Poirault-Chassac
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
,
Margot Rouaud
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
,
Hippolyte Guerineau
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
,
Blandine Dizier
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
,
Dominique Pidard
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
,
Pascale Gaussem**
1   Service d'hématologie biologique, AH-HP, Hopital Européen Georges Pompidou, Paris, France
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
,
Christilla Bachelot-Loza**
2   Université de Paris, Innovative Therapies in Haemostasis, INSERM U1140, Paris, France
› Author Affiliations
Funding This study received funding from the Institut National de la Santé et de la Recherche Médicale, Promex Stiftung für die Forschung Foundation, Université de Paris.

Abstract

Background Platelet cytosolic cyclic adenosine monophosphate (cAMP) levels are balanced by synthesis, degradation, and efflux. Efflux can occur via multidrug resistant protein-4 (MRP4; ABCC4) present on dense granule and/or plasma membranes. As lipid rafts have been shown to interfere on cAMP homeostasis, we evaluated the relationships between the distribution and activity of MRP4 in lipid rafts and cAMP efflux.

Methods Platelet activation and cAMP homeostasis were analyzed in human and wild-type or MRP4-deleted mouse platelets in the presence of methyl-β-cyclodextrin (MßCD) to disrupt lipid rafts, and of activators of the cAMP signalling pathways. Human platelet MRP4 and effector proteins of the cAMP pathway were analyzed by immunoblots in lipid rafts isolated by differential centrifugation.

Results MßCD dose dependently inhibited human and mouse platelet aggregation without affecting per se cAMP levels. An additive inhibitory effect existed between the adenylate cyclase (AC) activator forskolin and MßCD that was accompanied by an overincrease of cAMP, and which was significantly enhanced upon MRP4 deletion. Finally, an efflux of cAMP out of resting platelets incubated with prostaglandin E1 (PGE1) was observed that was partly dependent on MRP4. Lipid rafts contained a small fraction (≈15%) of MRP4 and most of the inhibitory G-protein Gi, whereas Gs protein, AC3, and phosphodiesterases PDE2 and PDE3A were all present as only trace amounts.

Conclusion Our results are in favour of part of MRP4 present at the platelet surface, including in lipid rafts. Lipid raft integrity is necessary for cAMP signalling regulation, although MRP4 and most players of cAMP homeostasis are essentially located outside rafts.

Authors' Contributions

T.B.R. conceived the study, designed and performed the research, analyzed data and wrote the manuscript. A.L. participated in the research, analyzed data, and participated in writing the initial and revised versions of the manuscript. A.M., B.D., F.M., S.P.C., M.R., H.G., B.D., and C.B.L. performed research and, together with DP, revised the manuscript and gave final approval. C.B.L. and P.G. conceived the study, designed the research, analyzed data and wrote the manuscript.


* T.B.R. and A.L. share co-first authorship.


** P.G. and C.B.L. share co-senior authorship.


Supplementary Material



Publication History

Received: 02 October 2020

Accepted: 12 April 2021

Accepted Manuscript online:
13 April 2021

Article published online:
25 June 2021

© 2021. Thieme. All rights reserved.

Georg Thieme Verlag KG
Rüdigerstraße 14, 70469 Stuttgart, Germany

 
  • References

  • 1 Smolenski A. Novel roles of cAMP/cGMP-dependent signaling in platelets. J Thromb Haemost 2012; 10 (02) 167-176
  • 2 Marcus A, Broekman M, Drosopoulos J. et al. Thromboregulation by endothelial cells: significance for occlusive vascular diseases. Arterioscler Thromb Vasc Biol 2001; 21 (02) 178-182
  • 3 Narumiya S, Sugimoto Y, Ushikubi F. Prostanoid receptors: structures, properties, and functions. Physiol Rev 1999; 79 (04) 1193-1226
  • 4 Burkhart JM, Vaudel M, Gambaryan S. et al. The first comprehensive and quantitative analysis of human platelet protein composition allows the comparative analysis of structural and functional pathways. Blood 2012; 120 (15) e73-e82
  • 5 Makhoul S, Walter E, Pagel O. et al. Effects of the NO/soluble guanylate cyclase/cGMP system on the functions of human platelets. Nitric Oxide 2018; 76: 71-80
  • 6 Taskén K, Aandahl EM. Localized effects of cAMP mediated by distinct routes of protein kinase A. Physiol Rev 2004; 84 (01) 137-167
  • 7 Nagy Z, Smolenski A. Cyclic nucleotide-dependent inhibitory signaling interweaves with activating pathways to determine platelet responses. Res Pract Thromb Haemost 2018; 2 (03) 558-571
  • 8 Hunter RW, Mackintosh C, Hers I. Protein kinase C-mediated phosphorylation and activation of PDE3A regulate cAMP levels in human platelets. J Biol Chem 2009; 284 (18) 12339-12348
  • 9 Stroop SD, Beavo JA. Structure and function studies of the cGMP-stimulated phosphodiesterase. J Biol Chem 1991; 266 (35) 23802-23809
  • 10 Turko IV, Francis SH, Corbin JD. Binding of cGMP to both allosteric sites of cGMP-binding cGMP-specific phosphodiesterase (PDE5) is required for its phosphorylation. Biochem J 1998; 329 (Pt 3): 505-510
  • 11 Hechler B, Gachet C. P2 receptors and platelet function. Purinergic Signal 2011; 7 (03) 293-303
  • 12 Gurbel PA, Tantry US. Combination antithrombotic therapies. Circulation 2010; 121 (04) 569-583
  • 13 Gresele P, Momi S, Falcinelli E. Anti-platelet therapy: phosphodiesterase inhibitors. Br J Clin Pharmacol 2011; 72 (04) 634-646
  • 14 Belleville-Rolland T, Sassi Y, Decouture B. et al. MRP4 (ABCC4) as a potential pharmacologic target for cardiovascular disease. Pharmacol Res 2016; 107: 381-389
  • 15 Cheepala SB, Pitre A, Fukuda Y. et al. The ABCC4 membrane transporter modulates platelet aggregation. Blood 2015; 126 (20) 2307-2319
  • 16 Russel FG, Koenderink JB, Masereeuw R. Multidrug resistance protein 4 (MRP4/ABCC4): a versatile efflux transporter for drugs and signalling molecules. Trends Pharmacol Sci 2008; 29 (04) 200-207
  • 17 Sassi Y, Abi-Gerges A, Fauconnier J. et al. Regulation of cAMP homeostasis by the efflux protein MRP4 in cardiac myocytes. FASEB J 2012; 26 (03) 1009-1017
  • 18 Borgognone A, Pulcinelli FM. Reduction of cAMP and cGMP inhibitory effects in human platelets by MRP4-mediated transport. Thromb Haemost 2012; 108 (05) 955-962
  • 19 Decouture B, Dreano E, Belleville-Rolland T. et al. Impaired platelet activation and cAMP homeostasis in MRP4-deficient mice. Blood 2015; 126 (15) 1823-1830
  • 20 Jedlitschky G, Tirschmann K, Lubenow LE. et al. The nucleotide transporter MRP4 (ABCC4) is highly expressed in human platelets and present in dense granules, indicating a role in mediator storage. Blood 2004; 104 (12) 3603-3610
  • 21 Jedlitschky G, Cattaneo M, Lubenow LE. et al. Role of MRP4 (ABCC4) in platelet adenine nucleotide-storage: evidence from patients with delta-storage pool deficiencies. Am J Pathol 2010; 176 (03) 1097-1103
  • 22 Mattiello T, Guerriero R, Lotti LV. et al. Aspirin extrusion from human platelets through multidrug resistance protein-4-mediated transport: evidence of a reduced drug action in patients after coronary artery bypass grafting. J Am Coll Cardiol 2011; 58 (07) 752-761
  • 23 Edwards HV, Christian F, Baillie GS. cAMP: novel concepts in compartmentalised signalling. Semin Cell Dev Biol 2012; 23 (02) 181-190
  • 24 Fischmeister R, Castro LR, Abi-Gerges A. et al. Compartmentation of cyclic nucleotide signaling in the heart: the role of cyclic nucleotide phosphodiesterases. Circ Res 2006; 99 (08) 816-828
  • 25 Zaccolo M, Pozzan T. Discrete microdomains with high concentration of cAMP in stimulated rat neonatal cardiac myocytes. Science 2002; 295 (5560): 1711-1715
  • 26 Esseltine JL, Scott JD. AKAP signaling complexes: pointing towards the next generation of therapeutic targets?. Trends Pharmacol Sci 2013; 34 (12) 648-655
  • 27 Raslan Z, Magwenzi S, Aburima A, Taskén K, Naseem K. Targeting of type I protein kinase A to lipid rafts is required for platelet inhibition by the 3′,5′-cyclic adenosine monophosphate-signaling pathway. J Thromb Haemost 2015; 13: 1721-1734
  • 28 Bodin S, Tronchère H, Payrastre B. Lipid rafts are critical membrane domains in blood platelet activation processes. Biochim Biophys Acta 2003; 1610 (02) 247-257
  • 29 Sezgin E, Levental I, Mayor S, Eggeling C. The mystery of membrane organization: composition, regulation and roles of lipid rafts. Nat Rev Mol Cell Biol 2017; 18 (06) 361-374
  • 30 Raslan Z, Naseem KM. Compartmentalisation of cAMP-dependent signalling in blood platelets: The role of lipid rafts and actin polymerisation. Platelets 2015; 26 (04) 349-357
  • 31 Zhang W, Trible RP, Samelson LE. LAT palmitoylation: its essential role in membrane microdomain targeting and tyrosine phosphorylation during T cell activation. Immunity 1998; 9 (02) 239-246
  • 32 Wonerow P, Obergfell A, Wilde JI. et al. Differential role of glycolipid-enriched membrane domains in glycoprotein VI- and integrin-mediated phospholipase Cgamma2 regulation in platelets. Biochem J 2002; 364 (Pt 3): 755-765
  • 33 Rabani V, Davani S, Gambert-Nicot S, Meneveau N, Montange D. Comparative lipidomics and proteomics analysis of platelet lipid rafts using different detergents. Platelets 2016; 27 (07) 634-641
  • 34 Nikolaev VO, Moshkov A, Lyon AR. et al. Beta2-adrenergic receptor redistribution in heart failure changes cAMP compartmentation. Science 2010; 327 (5973): 1653-1657
  • 35 Zidovetzki R, Levitan I. Use of cyclodextrins to manipulate plasma membrane cholesterol content: evidence, misconceptions and control strategies. Biochim Biophys Acta 2007; 1768 (06) 1311-1324
  • 36 Gousset K, Wolkers WF, Tsvetkova NM. et al. Evidence for a physiological role for membrane rafts in human platelets. J Cell Physiol 2002; 190 (01) 117-128
  • 37 Katsel PL, Tagliente TM, Schwarz TE, Craddock-Royal BD, Patel ND, Maayani S. Molecular and biochemical evidence for the presence of type III adenylyl cyclase in human platelets. Platelets 2003; 14 (01) 21-33
  • 38 Quinton TM, Kim S, Jin J, Kunapuli SP. Lipid rafts are required in Galpha(i) signaling downstream of the P2Y12 receptor during ADP-mediated platelet activation. J Thromb Haemost 2005; 3 (05) 1036-1041
  • 39 Raslan Z, Aburima A, Naseem KM. The spatiotemporal regulation of cAMP signaling in blood platelets—old friends and new players. Front Pharmacol 2015; 6: 266
  • 40 Mika D, Leroy J, Vandecasteele G, Fischmeister R. PDEs create local domains of cAMP signaling. J Mol Cell Cardiol 2012; 52 (02) 323-329
  • 41 Steinberg SF, Brunton LL. Compartmentation of G protein-coupled signaling pathways in cardiac myocytes. Annu Rev Pharmacol Toxicol 2001; 41: 751-773
  • 42 Burgers PP, Ma Y, Margarucci L. et al. A small novel A-kinase anchoring protein (AKAP) that localizes specifically protein kinase A-regulatory subunit I (PKA-RI) to the plasma membrane. J Biol Chem 2012; 287 (52) 43789-43797