Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Phosphatidylserine-dependent engulfment by macrophages of nuclei from erythroid precursor cells

Nature volume 437, pages 754–758 (2005)Cite this article

Abstract

Definitive erythropoiesis usually occurs in the bone marrow or fetal liver, where erythroblasts are associated with a central macrophage in anatomical units called ‘blood islands’1,2. Late in erythropoiesis, nuclei are expelled from the erythroid precursor cells and engulfed by the macrophages in the blood island2,3. Here we show that the nuclei are engulfed by macrophages only after they are disconnected from reticulocytes, and that phosphatidylserine, which is often used as an ‘eat me’ signal for apoptotic cells, is also used for the engulfment of nuclei expelled from erythroblasts. We investigated the mechanism behind the enucleation and engulfment processes by isolating late-stage erythroblasts from the spleens of phlebotomized mice. When these erythroblasts were cultured, the nuclei protruded spontaneously from the erythroblasts. A weak physical force could disconnect the nuclei from the reticulocytes. The released nuclei contained an undetectable level of ATP, and quickly exposed phosphatidylserine on their surface. Fetal liver macrophages efficiently engulfed the nuclei; masking the phosphatidylserine on the nuclei with the dominant-negative form of milk-fat-globule EGF8 (MFG-E8) prevented this engulfment.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Enucleation of erythroblasts.
Figure 2: Flow cytometric analysis of enucleation.
Figure 3: Exposure of PtdSer on nuclei released from erythroblasts.
Figure 4: Phosphatidylserine-dependent engulfment of nuclei.

Similar content being viewed by others

References

  1. Bessis, M., Mize, C. & Prenant, M. Erythropoiesis: comparison of in vivo and in vitro amplification. Blood Cells 4, 155–174 (1978)

    CAS  PubMed  Google Scholar 

  2. Sadahira, Y. & Mori, M. Role of the macrophage in erythropoiesis. Pathol. Int. 49, 841–848 (1999)

    Article  CAS  Google Scholar 

  3. Kawane, K. et al. Requirement of DNase II for definitive erythropoiesis in the mouse fetal liver. Science 292, 1546–1549 (2001)

    Article  ADS  CAS  Google Scholar 

  4. Pannacciulli, I. M. et al. Effect of bleeding on in vivo and in vitro colony-forming hemopoietic cells. Acta Haematol. 58, 27–33 (1977)

    Article  CAS  Google Scholar 

  5. Zhang, D. et al. An optimized system for studies of EPO-dependent murine pro-erythroblast development. Exp. Hematol. 29, 1278–1288 (2001)

    Article  CAS  Google Scholar 

  6. Socolovsky, M. et al. Ineffective erythropoiesis in Stat5a-/-5b-/- mice due to decreased survival of early erythroblasts. Blood 98, 3261–3273 (2001)

    Article  CAS  Google Scholar 

  7. Repasky, E. A. & Eckert, B. S. The effect of cytochalasin B on the enucleation of erythroid cells in vitro. Cell Tissue Res. 221, 85–91 (1981)

    Article  CAS  Google Scholar 

  8. Wu, H., Liu, X., Jaenisch, R. & Lodish, H. F. Generation of committed erythroid BFU-E and CFU-E progenitors does not require erythropoietin or the erythropoietin receptor. Cell 83, 59–67 (1995)

    Article  CAS  Google Scholar 

  9. Broudy, V. C., Lin, N., Brice, M., Nakamoto, B. & Papayannopoulou, T. Erythropoietin receptor characteristics on primary human erythroid cells. Blood 77, 2583–2590 (1991)

    CAS  PubMed  Google Scholar 

  10. Carlile, G. W., Smith, D. H. & Wiedmann, M. Caspase-3 has a nonapoptotic function in erythroid maturation. Blood 103, 4310–4316 (2004)

    Article  CAS  Google Scholar 

  11. Zermati, Y. et al. Caspase activation is required for terminal erythroid differentiation. J. Exp. Med. 193, 247–254 (2001)

    Article  CAS  Google Scholar 

  12. Fadok, V. A., Bratton, D. L., Frasch, S. C., Warner, M. L. & Henson, P. M. The role of phosphatidylserine in recognition of apoptotic cells by phagocytes. Cell Death Differ. 5, 551–562 (1998)

    Article  CAS  Google Scholar 

  13. Balasubramanian, K. & Schroit, A. J. Aminophospholipid asymmetry: A matter of life and death. Annu. Rev. Physiol. 65, 701–734 (2003)

    Article  CAS  Google Scholar 

  14. Kawane, K. et al. Impaired thymic development in mouse embryos deficient in apoptotic DNA degradation. Nature Immunol. 4, 138–144 (2003)

    Article  CAS  Google Scholar 

  15. Hanayama, R. et al. Identification of a factor that links apoptotic cells to phagocytes. Nature 417, 182–187 (2002)

    Article  ADS  CAS  Google Scholar 

  16. Hanspal, M. Importance of cell–cell interactions in regulation of erythropoiesis. Curr. Opin. Hematol. 4, 142–147 (1997)

    Article  CAS  Google Scholar 

  17. Davies, P. F. Flow-mediated endothelial mechanotransduction. Physiol. Rev. 75, 519–560 (1995)

    Article  CAS  Google Scholar 

  18. Schlegel, R. A., Phelps, B. M., Waggoner, A., Terada, L. & Williamson, P. Binding of merocyanine 540 to normal and leukemic erythroid cells. Cell 20, 321–328 (1980)

    Article  CAS  Google Scholar 

  19. Geiduschek, J. B. & Singer, S. J. Molecular changes in the membranes of mouse erythroid cells accompanying differentiation. Cell 16, 149–163 (1979)

    Article  CAS  Google Scholar 

  20. Strehler, E. E. & Treiman, M. Calcium pumps of plasma membrane and cell interior. Curr. Mol. Med. 4, 323–335 (2004)

    Article  CAS  Google Scholar 

  21. Tang, X., Halleck, M. S., Schlegel, R. A. & Williamson, P. A subfamily of P-type ATPases with aminophospholipid transporting activity. Science 272, 1495–1497 (1996)

    Article  ADS  CAS  Google Scholar 

  22. Zhou, Q. et al. Molecular cloning of human plasma membrane phospholipid scramblase. A protein mediating transbilayer movement of plasma membrane phospholipids. J. Biol. Chem. 272, 18240–18244 (1997)

    Article  CAS  Google Scholar 

  23. Zhou, Q., Zhao, J., Wiedmer, T. & Sims, P. J. Normal hemostasis but defective hematopoietic response to growth factors in mice deficient in phospholipid scramblase 1. Blood 99, 4030–4038 (2002)

    Article  CAS  Google Scholar 

  24. Williamson, P. & Schlegel, R. A. Hide and seek: the secret identity of the phosphatidylserine receptor. J. Biol. 3, 14 (2004)

    Article  Google Scholar 

  25. Krahling, S., Callahan, M. K., Williamson, P. & Schlegel, R. A. Exposure of phosphatidylserine is a general feature in the phagocytosis of apoptotic lymphocytes by macrophages. Cell Death Differ. 6, 183–189 (1999)

    Article  CAS  Google Scholar 

  26. Fadok, V. A. et al. A receptor for phosphatidylserine-specific clearance of apoptotic cells. Nature 405, 85–90 (2000)

    Article  ADS  CAS  Google Scholar 

  27. Kawasaki, Y., Nakagawa, A., Nagaosa, K., Shiratsuchi, A. & Nakanishi, Y. Phosphatidylserine binding of class B scavenger receptor type I, a phagocytosis receptor of testicular Sertoli cells. J. Biol. Chem. 277, 27559–27566 (2002)

    Article  CAS  Google Scholar 

  28. Hanayama, R., Tanaka, M., Miwa, K. & Nagata, S. Expression of developmental endothelial locus-1 in a subset of macrophages for engulfment of apoptotic cells. J. Immunol. 172, 3876–3882 (2004)

    Article  CAS  Google Scholar 

  29. Asano, K. et al. Masking of phosphatidylserine inhibits apoptotic cell engulfment and induces autoantibody production in mice. J. Exp. Med. 200, 459–467 (2004)

    Article  CAS  Google Scholar 

  30. Takeshita, S., Kaji, K. & Kudo, A. Identification and characterization of the new osteoclast progenitor with macrophage phenotypes being able to differentiate into mature osteoclasts. J. Bone Miner. Res. 15, 1477–1488 (2000)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank H. Fukuyama for help at the initial stage of this work; H. Suzuki, I. Ito and S. Hanasaki for taking the time-lapse video; T. Yanagida for discussion; and M. Fujii and M. Harayama for secretarial assistance. This work was supported in part by Grants-in-Aid from the Ministry of Education, Science, Sports and Culture in Japan.Author Contributions H.Y. performed most of the experiments. M.K. carried out the electron microscopy analysis, and Y.M. took the time-lapse video. K.K., Y.U. and S.N. contributed to the data analysis. S.N. was responsible for planning the project.

Author information

Authors and Affiliations

  1. Department of Genetics, Osaka University, 565-0871, Osaka, Japan

    Hideyuki Yoshida, Kohki Kawane & Shigekazu Nagata

  2. Department of Cell Biology and Neuroscience, Graduate School of Medicine, Osaka University, 565-0871, Osaka, Japan

    Masato Koike & Yasuo Uchiyama

  3. Laboratory of Genetics, Integrated Biology Laboratories, Graduate School of Frontier Biosciences, Osaka University, Osaka, 565-0871, Japan

    Kohki Kawane & Shigekazu Nagata

  4. Core Research for Evolutional Science and Technology, Japan Science and Technology Corporation, 565-0871, Osaka, Japan

    Kohki Kawane & Shigekazu Nagata

  5. Sakura Motion Picture Co. Ltd, 20-1 Sendagaya 4-chome, Shibuya, Tokyo, 151-0051, Japan

    Yoshimi Mori

Authors
  1. Hideyuki Yoshida
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Kohki Kawane
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Masato Koike
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yoshimi Mori
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Yasuo Uchiyama
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Shigekazu Nagata
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Shigekazu Nagata.

Ethics declarations

Competing interests

Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

Supplementary information

Supplementary Video S1

Time-lapse video of the enuclation of an erythroblast. Erythroblasts (1 X 106 cells/ml) were incubated at 37°C in the presence of erythropoietin and transferin. The morphological change of one erythroblast was followed for 3 h, and is reproduced in a 10-s video. (MOV 947 kb)

Supplementary Video S2

Time-lapse video of the engulfment of nuclei by macrophages. Fetal liver macrophages (2 x 104 cells) from DNase II-/- embryos were co-cultured at 37°C with 5 x 105 nuclei isolated from erythroblasts in the absence of 0.5 µg/ml of the D89E mutant of MFG-E8. The engulfment of nuclei by one macrophage was followed for 3 h, and reproduced in a 10-s video. At the end of the incubation, the cells were stained with SYTOX to visualize the engulfed nuclei. (MOV 664 kb)

Supplementary Video S3

Time-lapse videos of the engulfment of nuclei by macrophages. Fetal liver macrophages (2 x 104 cells) from DNase II-/- embryos were co-cultured at 37°C with 5 x 105 nuclei isolated from erythroblasts in the presence of 0.5 µg/ml of the D89E mutant of MFG-E8. The engulfment of nuclei by one macrophage was followed for 3 h, and reproduced in a 10-s video. At the end of the incubation, the cells were stained with SYTOX to visualize the engulfed nuclei. (MOV 637 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Yoshida, H., Kawane, K., Koike, M. et al. Phosphatidylserine-dependent engulfment by macrophages of nuclei from erythroid precursor cells. Nature 437, 754–758 (2005). https://doi.org/10.1038/nature03964

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature03964

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing