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Association of Gpx1 fluctuation in cell cycle progression

  • Khudishta Aktar
  • Abdul Kafi
  • Ravinder DahiyaEmail author
Article

Abstract

This research demonstrates fluctuation of glutathione peroxidase1 (Gpx1) throughout cell cycle progression with significant decreased expression at mitosis of HeLa cell. This was achieved with western blot (WB) analysis of target proteins from each phase of synchronized cells. The synchronizations were performed with double thymidine (T/T) for G1/S arrest and thymidine followed by nocodazole (T/N) for G2/M arrest. The G1/S arrested cells were released in fresh medium for 3, 6, 9, 10, and 15h to obtain cell at each phase such as gap1 (G1), synthesis (S), gap2 (G2), mitosis (M), and gap1 (G1) phase, respectively, for investigating Gpx1 expression throughout a complete cycle. The synchronizations were confirmed using fluorescence activated cell sorting (FACS) and WB analysis of phase-specific markers. The fluctuations of Gpx1 expression were verified with universal protein actin and peroxiredoxin1 (Prx1) which are stable throughout the cell cycle. Intriguingly, immunoblots showed the level of Gpx1 decreases at mitosis phase and increased during mitotic exit to G1 phase in HeLa cells, while Prx1 protein level remained constant. The fractionation experiments reveal that only the cytosolic Gpx1 was decreased while their levels at mitochondria remain constant. The highest levels of mitochondrial ROS were measured in mitosis phase with FACS analysis using Mito sox indicating that antioxidant activity of Gpx1 for detoxifying excessive induced endogenous reactive oxygen species (ROS) in the mitosis phase could be the reason for such decreasing level. For unfolding the molecular mechanism of such decreased expression, the Gpx1 was investigated at transcriptional, translational, and proteosomal level. The results revealed that translational mechanism is involve in the decreased expression rather than transcriptional or proteosomal degradation at mitosis phase. This finding supports that Gpx1 is involved in the cell cycle progression through regulation of endogenous ROS. Based on this observation, further research could uncover their possible association with the infinitive division of a cancer cell.

Keywords

Glutathione peroxidase Cell division Mitosis Reactive oxygen species Antioxidant protein 

Notes

Authors’ contributions

M.K.A. was involved in designing and performing the research work; M.K.A, M.A.K, and R.D. were involved writing and formatting the manuscript. All the authors reviewed the manuscript.

Funding information

This research was financially funded by the International Exchange Fellowship grant, Brain Korea 21 Fellowship grant, and Marie Curie IF grant.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

References

  1. Barrera G (2012) Oxidative stress and lipid peroxidation products in cancer progression and therapy. ISRN Oncol 137289:1–21.  https://doi.org/10.5402/2012/137289 Google Scholar
  2. Boonstra J, Post JA (2004) Molecular events associated with reactive oxygen species and cell cycle progression in mammalian cells. Gene 337:1–13CrossRefGoogle Scholar
  3. Borchert A, Wang CC, Ufer C, Schiebel H, Savaskan NE, Kuhn H (2006) The role of phospholipid hydroperoxide glutathione peroxidase isoforms in murine embryogenesis. J Biol Chem 281:19655–19664CrossRefGoogle Scholar
  4. Circu ML, Aw TY (2010) Reactive oxygen species, cellular redox systems and apoptosis. Free Radic Biol Med 48:749–762CrossRefGoogle Scholar
  5. de-Haan JB, Cooper ME (2011) Targeted antioxidant therapies in hyperglycemia-mediated endothelial dysfunction. Front Biosci 3:709–729CrossRefGoogle Scholar
  6. Diamond AM (2015) The subcellular location of selenoproteins and the impact on their function. Nutrients 7:3938–3948CrossRefGoogle Scholar
  7. Espinosa-Diez C, Miguel V, Mennerich D, Kietzmann T, Sánchez-Pérez P, Cadenas S, Lamas S (2015) Antioxidant responses and cellular adjustments to oxidative stress. Redox Biol 6:183–197CrossRefGoogle Scholar
  8. Fang G, Yu H, Kirschner MW (1998) Direct binding of CDC20 protein family members activates the anaphase-promoting complex in mitosis and G1. Mol Cell 2:163–171CrossRefGoogle Scholar
  9. Gavet O, Pines J (2010) Progressive activation of CyclinB1-Cdk1 coordinates entry to mitosis. Dev Cell 18:533–543CrossRefGoogle Scholar
  10. Goldson AJ, Fairweather-Tait SJ, Armah CN, Bao Y, Broadley MR, Dainty JR, Furniss C, Hart DJ, Teucher B, Hurst R (2011) Effects of selenium supplementation on Selenoprotein gene expression and response to influenza vaccine challenge: a randomised controlled trial. PLoS One 6:e14771.  https://doi.org/10.1371/journal.pone.0014771 CrossRefGoogle Scholar
  11. Han Y, Ishibashi S, Iglesias-Gonzalez J, Chen Y, Love NR, Amaya E (2018) Ca2+-induced mitochondrial ROS regulate the early embryonic cell cycle. Cell Rep 22:218–231CrossRefGoogle Scholar
  12. Handy DE, Lubos E, Yang Y, Galbraith JD, Kelly N, Zhang Y-Y, Leopold JA, Loscalzo J (2009) Glutathione peroxidase-1 regulates mitochondrial function to modulate redox-dependent cellular responses. J Biol Chem 284:11913–11921.  https://doi.org/10.1074/jbc.M900392200 CrossRefGoogle Scholar
  13. Havens CG, Ho A, Yoshioka N, Dowdy SF (2006) Regulation of late G1/S phase transition and APCCdh1 by reactive oxygen species. Mol Cell Biol 26:4701–4711CrossRefGoogle Scholar
  14. Herbette S, Roeckel-Drevet P, Drevet JR (2007) Seleno-independent glutathione peroxidases. More than simple antioxidant scavengers. FEBS J 274:2163–2180CrossRefGoogle Scholar
  15. Hugo M, Korovila I, Köhlera M, García-García C, Cabrera-García JD, Marina A, Martínez-Ruiz A, Grune T (2018) Early cysteine-dependent inactivation of 26S proteasomes does not involve particle disassembly. Redox Biol 16:123–128CrossRefGoogle Scholar
  16. Ibañez IL, Policastro LL, Tropper I, Bracalente C, Almieri MA, Rojas PA, Molinari BL, Durán H (2011) H2O2 scavenging inhibits G1/S transition by increasing nuclear levels of p27KIP1. Cancer Lett 305:58–68CrossRefGoogle Scholar
  17. Idelchik MPS, Begley U, Begley TJ, Melendez JA (2017) Mitochondrial ROS control of cancer. Semin Cancer Biol 47:57–66CrossRefGoogle Scholar
  18. Ighodaro OM, Akinloye OA (2017) First line defence antioxidants-superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX): their fundamental role in the entire antioxidant defence grid. J Med 54:287–293.  https://doi.org/10.1016/j.ajme.2017.09.001 Google Scholar
  19. Kafi MA, Cho HY, Choi JW (2016) Engineered peptide-based nanobiomaterials for electrochemical cell chip. Nano Convergence 3(17):17.  https://doi.org/10.1186/s40580-016-0077-7 CrossRefGoogle Scholar
  20. Kafi MA, El-Said WA, Kim TH, Choi JW (2012) Cell adhesion, spreading, and proliferation on surface functionalized with RGD nanopillar arrays. Biomaterials 33:731–739CrossRefGoogle Scholar
  21. Kafi MA, Kim TH, An JH, Choi JW (2011) Fabrication of cell chip for detection of cell cycle progression based on electrochemical method. Anal Chem 83:2104–2111CrossRefGoogle Scholar
  22. Kryukov GV, Castellano S, Novoselov SV, Lobanov AV, Zehtab O, Guigo R, Gladyshev VN (2003) Characterization of mammalian selenoproteomes. Science 300:1439–1443CrossRefGoogle Scholar
  23. Li H, Horke S, Förstermann U (2013b) Oxidative stress in vascular disease and its pharmacological prevention. Trends Pharmacol Sci 34:313319CrossRefGoogle Scholar
  24. Li R, Jen N, Yu F, Hsiai TK (2011) Assessing mitochondrial redox status by flow cytometric methods: vascular response to fluid shear stress. Curr Protoc Cytom Chapter 9.  https://doi.org/10.1002/0471142956.cy0937s58
  25. Li X, Fang P, Mai J, Choi ET, Wang H, Yang XF (2013a) Targeting mitochondrial reactive oxygen species as novel therapy for inflammatory diseases and cancers. J Hematol Oncol 6:19CrossRefGoogle Scholar
  26. Lu J, Holmgren A (2009) Selenoproteins. J Biol Chem 284:723–727CrossRefGoogle Scholar
  27. Lubos E, Loscalzo J, Handy DE (2011) Glutathione peroxidase-1 in health and disease: from molecular mechanisms to therapeutic opportunities. Antioxid Redox Signal 15:1957–1997CrossRefGoogle Scholar
  28. Ludke A, Akolkar G, Ayyappan P, Sharma AK, Singal PK (2017) Time course of changes in oxidative stress and stress-induced proteins in cardiomyocytes exposed to doxorubicin and prevention by vitamin C. PLoS One 12:e0179452.  https://doi.org/10.1371/journal.pone.0179452
  29. Mehdi Y, Dufrasne I (2016) Selenium in cattle: a review. Molecules 21:545.  https://doi.org/10.3390/molecules21040545 CrossRefGoogle Scholar
  30. Neve J (1995) Human selenium supplementation as assessed by changes in blood selenium concentration and glutathione peroxidase activity. J Trace Elem Med Biol 9:65–73CrossRefGoogle Scholar
  31. Nielsen PS, Riber-Hansen R, Jensen TO, Schmidt H, Steiniche T (2013) Proliferation indices of phosphohistone H3 and Ki67: strong prognostic markers in a consecutive cohort with stage I/II melanoma. Mod Pathol 26:404–413CrossRefGoogle Scholar
  32. Onumah OE, Jules GE, Zhao Y, Zhou L, Yang H, Guo Z (2009) Overexpression of catalase delays G0/G1 to S phase transition during cell cycle progression in mouse aortic endothelial cell. Free Radic Biol Med 46:1658–1667CrossRefGoogle Scholar
  33. Power SK, Jackson MJ (2008) Exercise-induced oxidative stress: cellular mechanisms and impact on muscle force production. Physiol Rev 88:1243–1276CrossRefGoogle Scholar
  34. Raha S, McEachern GE, Myint AT, Robinson BH (2000) Superoxides from mitochondrial complex III: the ole of manganese superoxide dismutase. Free Radic Biol Med 29:170–180CrossRefGoogle Scholar
  35. Ray PD, Huang BW, Tsuji Y (2012) Reactive oxygen species (ROS) homeostasis and redox regulation in cellular signalling. Cell Signal 24:981–990.  https://doi.org/10.1016/j.cellsig.2012.01.008 CrossRefGoogle Scholar
  36. Redza-Dutordoir M, Averill-Bates DA (2016) Activation of apoptosis signalling pathways by reactive oxygen species. Biochim Biophys Acta 1863:2977–2992CrossRefGoogle Scholar
  37. Schorl C, Sedivy JM (2007) Analysis of cell cycle phases and progression in cultured mammalian cells. Methods Methods 41:143–150CrossRefGoogle Scholar
  38. Speckmann B, Bidmon HJ, Pinto A, Anlauf M, Sies H Steinbrenner H (2011) Induction of glutathione peroxidase 4 expression during enterocytic cell differentiation. J Biol Chem 286:10764–10772CrossRefGoogle Scholar
  39. Wang K, Zhang T, Dong Q, Nice EC, Huang C, Wei Y (2013) Redox homeostasis: the linchpin in stem cell self-renewal and differentiation. Cell Death Differ 537:e537.  https://doi.org/10.1038/cddis.2013.50 CrossRefGoogle Scholar
  40. Waris G, Ahsan H (2006) Reactive oxygen species: role in the development of cancer and various chronic conditions. J Carcinog 5:14.  https://doi.org/10.1186/1477-3163-5-14 CrossRefGoogle Scholar
  41. Wellen KE, Thompson CB (2010) Cellular metabolic stress: considering how cells respond to nutrient excess. Mol Cell 40:323–332CrossRefGoogle Scholar
  42. You J, Chan Z (2015) ROS regulation during abiotic stress responses in crop plants. Front Plant Sci 6:1092CrossRefGoogle Scholar

Copyright information

© The Society for In Vitro Biology 2019

Authors and Affiliations

  1. 1.BEST group, School of EngineeringUniversity of GlasgowGlasgowUK
  2. 2.Department of Life ScienceEwha Womans UniversitySeoulSouth Korea

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