Thromb Haemost 2015; 114(05): 969-981
DOI: 10.1160/TH14-09-0727
Cellular Haemostasis and Platelets
Schattauer GmbH

Role of mTOR1 and mTOR2 complexes in MEG-01 cell physiology

Esther López
1   Department of Physiology (PHYCELL), University of Extremadura, Cáceres, Spain
,
Alejandro Berna-Erro
1   Department of Physiology (PHYCELL), University of Extremadura, Cáceres, Spain
,
Javier J. López
1   Department of Physiology (PHYCELL), University of Extremadura, Cáceres, Spain
,
María P. Granados
1   Department of Physiology (PHYCELL), University of Extremadura, Cáceres, Spain
,
Nuria Bermejo
2   Hematology Unit, San Pedro de Alcántara Hospital, Cáceres, Spain
,
José M. Brull
3   Blood donation centre Extremadura, Mérida, Spain
,
Ginés M. Salido
1   Department of Physiology (PHYCELL), University of Extremadura, Cáceres, Spain
,
Juan A. Rosado
1   Department of Physiology (PHYCELL), University of Extremadura, Cáceres, Spain
,
Pedro C. Redondo
1   Department of Physiology (PHYCELL), University of Extremadura, Cáceres, Spain
› Institutsangaben
Weitere Informationen

Publikationsverlauf

Received: 04. September 2014

Accepted after major revision: 23. Mai 2015

Publikationsdatum:
06. Dezember 2017 (online)

Summary

The function of the mammalian target of rapamycin (mTOR) is upregulated in response to cell stimulation with growing and differentiating factors. Active mTOR controls cell proliferation, differentiation and death. Since mTOR associates with different proteins to form two functional macromolecular complexes, we aimed to investigate the role of the mTORI and mTOR2 complexes in MEG-01 cell physiology in response to thrombopoietin (TPO). By using mTOR antagonists and overexpressing FKBP38, we have explored the role of both mTOR complexes in proliferation, apoptosis, maturation-like mechanisms, endoplasmic reticulum-stress and the intracellular location of both active mTOR complexes during MEG-01 cell stimulation with TPO. The results demonstrate that mTOR1 and mTOR2 complexes play different roles in the physiology of MEG-01 cells and in the maturation-like mechanisms; hence, these findings might help to understand the mechanism underlying generation of platelets.

 
  • References

  • 1 Jiang Y. mTOR goes to the nucleus. Cell Cycle 2010; 9: 868.
  • 2 Sarbassov DD, Ali SM, Sengupta S. et al. Prolonged rapamycin treatment inhibits mTORC2 assembly and Akt/PKB. Mol Cell 2006; 22: 159-168.
  • 3 Akcakanat A, Singh G, Hung MC. et al. Rapamycin regulates the phosphorylation of rictor. Biochem Biophys Res Commun 2007; 362: 330-333.
  • 4 Thoreen CC, Kang SA, Chang JW. et al. An ATP-competitive mammalian target of rapamycin inhibitor reveals rapamycin-resistant functions of mTORC1. J Biol Chem 2009; 284: 8023-8032.
  • 5 Liu Q, Kang SA, Thoreen CC. et al. Development of ATP-competitive mTOR inhibitors. Methods Mol Biol 2012; 821: 447-460.
  • 6 Thoreen CC, Sabatini DM. Rapamycin inhibits mTORC1, but not completely. Autophagy 2009; 5: 725-726.
  • 7 Lopez E, Berna-Erro A, Bermejo N. et al. Long-term mTOR inhibitors administration evokes altered calcium homeostasis and platelet dysfunction in kidney transplant patients. J Cell Mol Med 2013; 17: 636-647.
  • 8 Murgia MG, Jordan S, Kahan BD. The side effect profile of sirolimus: a phase I study in quiescent cyclosporine-prednisone-treated renal transplant patients. Kidney Int 1996; 49: 209-216.
  • 9 Kelly PA, Gruber SA, Behbod F. et al. Sirolimus, a new, potent immunosuppressive agent. Pharmacotherapy 1997; 17: 1148-1156.
  • 10 Kahan BD. The potential role of rapamycin in pediatric transplantation as observed from adult studies. Pediatr Transplant 1999; 3: 175-180.
  • 11 Lopez JJ, Palazzo A, Chaabane C. et al. Crucial role for endoplasmic reticulum stress during megakaryocyte maturation. Arterioscler Thromb Vasc Biol 2013; 33: 2750-2758.
  • 12 Schipper LF, Brand A, Fibbe WE. et al. Functional characterisation of TPO-expanded CD34+ cord blood cells identifies CD34-CD61-cells as platelet-producing cells early after transplantation in NOD/SCID mice and rCD34+ cells as CAFC colony-forming cells. Stem Cells 2012; 30: 988-996.
  • 13 Battinelli E, Willoughby SR, Foxall T. et al. Induction of platelet formation from megakaryocytoid cells by nitric oxide. Proc Natl Acad Sci USA 2001; 98: 14458-14463.
  • 14 Erhardt JA, Erickson-Miller CL, Aivado M. et al. Comparative analyses of the small molecule thrombopoietin receptor agonist eltrombopag and thrombopoietin on in vitro platelet function. Exp Hematol 2009; 37: 1030-1037.
  • 15 Sun H, Tsai Y, Nowak I. et al. Eltrombopag, a thrombopoietin receptor agonist, enhances human umbilical cord blood hematopoietic stem/primitive progenitor cell expansion and promotes multi-lineage hematopoiesis. Stem Cell Res 2012; 9: 77-86.
  • 16 Raslova H, Baccini V, Loussaief L. et al. Mammalian target of rapamycin (mTOR) regulates both proliferation of megakaryocyte progenitors and late stages of megakaryocyte differentiation. Blood 2006; 107: 2303-2310.
  • 17 Rank A, Nieuwland R, Delker R. et al. Surveillance of megakaryocyte function by measurement of CD61-exposing microparticles in allogeneic hematopoietic stem cell recipients. Clin Transplant 2011; 25: E233-242.
  • 18 Ojima Y, Duncan MT, Nurhayati RW. et al. Synergistic effect of hydrogen peroxide on polyploidisation during the megakaryocytic differentiation of K562 leukemia cells by PMA. Exp Cell Res 2013; 319: 2205-2215.
  • 19 Li J, Kuter DJ. The end is just the beginning: megakaryocyte apoptosis and platelet release. Int J Hematol 2001; 74: 365-374.
  • 20 Xiao Y, Zheng Y, Tan P. et al. Overexpression of nuclear distribution protein (hNUDC) causes pro-apoptosis and differentiation in Dami megakaryocytes. Cell Prolif 2013; 46: 576-585.
  • 21 Yoon MS, Sun Y, Arauz E. et al. Phosphatidic acid activates mammalian target of rapamycin complex 1 (mTORC1) kinase by displacing FK506 binding protein 38 (FKBP38) and exerting an allosteric effect. J Biol Chem 2011; 286: 29568-29574.
  • 22 Bai X, Ma D, Liu A. et al. Rheb activates mTOR by antagonizing its endogenous inhibitor, FKBP38. Science 2007; 318: 977-980.
  • 23 Ozoren N, El-Deiry WS. Cell surface Death Receptor signaling in normal and cancer cells. Semin Cancer Biol 2003; 13: 135-147.
  • 24 Betz C, Stracka D, Prescianotto-Baschong C. et al. Feature Article: mTOR complex 2-Akt signaling at mitochondria-associated endoplasmic reticulum membranes (MAM) regulates mitochondrial physiology. Proc Natl Acad Sci USA 2013; 110: 12526-12534.
  • 25 Wang ZQ, Yang Y, Lu T. et al. Protective effect of autophagy inhibition on ischemia-reperfusion-induced injury of N2a cells. J Huazhong Univ Sci Technolog Med Sci 2013; 33: 810-816.
  • 26 Mizushima N, Yoshimori T. How to interpret LC3 immunoblotting. Autophagy 2007; 3: 542-545.
  • 27 Guerriero R, Parolini I, Testa U. et al. Inhibition of TPO-induced MEK or mTOR activity induces opposite effects on the ploidy of human differentiating megakaryocytes. J Cell Sci 2006; 119: 744-752.
  • 28 Saffak T, Schafer S, Haas C. et al. Regulation of the human thromboxane A2 receptor gene in human megakaryoblastic MEG-01 cells. Prostaglandins Leukot Essent Fatty Acids 2003; 69: 299-306.
  • 29 Sattler M, Durstin MA, Frank DA. et al. The thrombopoietin receptor c-MPL activates JAK2 and TYK2 tyrosine kinases. Exp Hematol 1995; 23: 1040-1048.
  • 30 Sattler M, Salgia R, Durstin MA. et al. Thrombopoietin induces activation of the phosphatidylinositol-3' kinase pathway and formation of a complex containing p85PI3K and the protooncoprotein p120CBL. J Cell Physiol 1997; 171: 28-33.
  • 31 Liu ZJ, Italiano Jr. J, Ferrer-Marin F. et al. Developmental differences in megakaryocytopoiesis are associated with up-regulated TPO signaling through mTOR and elevated GATA-1 levels in neonatal megakaryocytes. Blood 2011; 117: 4106-4117.
  • 32 Hong JC, Kahan BD. Sirolimus-induced thrombocytopenia and leukopenia in renal transplant recipients: risk factors, incidence, progression, and management. Transplantation 2000; 69: 2085-2090.
  • 33 Shirane M, Nakayama KI. Inherent calcineurin inhibitor FKBP38 targets Bcl-2 to mitochondria and inhibits apoptosis. Nat Cell Biol 2003; 5: 28-37.
  • 34 Shirane-Kitsuji M, Nakayama KI. Mitochondria: FKBP38 and mitochondrial degradation. Int J Biochem Cell Biol 2014; 51: 19-22.
  • 35 Zou H, Lai Y, Zhao X. et al. Regulation of mammalian target of rapamycin complex 1 by Bcl-2 and Bcl-XL proteins. J Biol Chem 2013; 288: 28824-28830.
  • 36 Huang AC, Lien JC, Lin MW. et al. Tetrandrine induces cell death in SAS human oral cancer cells through caspase activation-dependent apoptosis and LC3-I and LC3-II activation-dependent autophagy. Int J Oncol 2013; 43: 485-494.
  • 37 Nakamura O, Hitora T, Akisue T. et al. Inhibition of induced autophagy increases apoptosis of Nara-H cells. Int J Oncol 2011; 39: 1545-1552.
  • 38 Diaz-Troya S, Florencio FJ, Crespo JL. Target of rapamycin and LST8 proteins associate with membranes from the endoplasmic reticulum in the unicellular green alga Chlamydomonas reinhardtii. Eukaryot Cell 2008; 7: 212-222.
  • 39 Desai BN, Myers BR, Schreiber SL. FKBP12-rapamycin-associated protein associates with mitochondria and senses osmotic stress via mitochondrial dysfunction. Proc Natl Acad Sci USA 2002; 99: 4319-4324.