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.

  • Article
  • Published:

Death-receptor O-glycosylation controls tumor-cell sensitivity to the proapoptotic ligand Apo2L/TRAIL

Abstract

Apo2L/TRAIL stimulates cancer cell death through the proapoptotic receptors DR4 and DR5, but the determinants of tumor susceptibility to this ligand are not fully defined. mRNA expression of the peptidyl O-glycosyltransferase GALNT14 correlated with Apo2L/TRAIL sensitivity in pancreatic carcinoma, non–small-cell lung carcinoma and melanoma cell lines, and up to 30% of samples from various human malignancies showed GALNT14 overexpression. RNA interference of GALNT14 reduced cellular Apo2L/TRAIL sensitivity, whereas overexpression increased responsiveness. Biochemical analysis of DR5 identified several ectodomain O-(N-acetyl galactosamine–galactose–sialic acid) structures. Sequence comparison predicted conserved extracellular DR4 and DR5 O-glycosylation sites; progressive mutation of the DR5 sites attenuated apoptotic signaling. O-glycosylation promoted ligand-stimulated clustering of DR4 and DR5, which mediated recruitment and activation of the apoptosis-initiating protease caspase-8. These results uncover a new link between death-receptor O-glycosylation and apoptotic signaling, providing potential predictive biomarkers for Apo2L/TRAIL-based cancer therapy.

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

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

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

Figure 1: Specific O-glycosylation enzyme expression correlates with sensitivity to Apo2L/TRAIL.
Figure 2: Inhibition of O-glycosylation enzymes reduces sensitivity to Apo2L/TRAIL.
Figure 3: Overexpression of O-glycosylation enzymes promotes sensitivity to Apo2L/TRAIL.
Figure 4: Modulation of O-glycosylation enzymes affects caspase-8 activation.
Figure 5: Identification of potential O-glycosylation sites in the ectodomain of DR4 and DR5.
Figure 6: Inhibition of O-glycosylation impairs Apo2L/TRAIL-induced receptor clustering.

Similar content being viewed by others

Accession codes

Accessions

Gene Expression Omnibus

References

  1. Danial, N.N. & Korsmeyer, S.J. Cell death: critical control points. Cell 116, 205–219 (2004).

    Article  CAS  Google Scholar 

  2. Fesik, S.W. Promoting apoptosis as a strategy for cancer drug discovery. Nat. Rev. Cancer 5, 876–885 (2005).

    Article  CAS  Google Scholar 

  3. Dalton, W.S. & Friend, S.H. Cancer biomarkers—an invitation to the table. Science 312, 1165–1168 (2006).

    Article  CAS  Google Scholar 

  4. Ashkenazi, A. & Dixit, V.M. Death receptors: signaling and modulation. Science 281, 1305–1308 (1998).

    Article  CAS  Google Scholar 

  5. Ashkenazi, A. Targeting death and decoy receptors of the tumour-necrosis factor superfamily. Nat. Rev. Cancer 2, 420–430 (2002).

    Article  CAS  Google Scholar 

  6. Kischkel, F.C. et al. Cytotoxicity-dependent APO-1 (Fas/CD95)-associated proteins form a death-inducing signaling complex (DISC) with the receptor. EMBO J. 14, 5579–5588 (1995).

    Article  CAS  Google Scholar 

  7. Kischkel, F.C. et al. Apo2L/TRAIL-dependent recruitment of endogenous FADD and caspase-8 to death receptors 4 and 5. Immunity 12, 611–620 (2000).

    Article  CAS  Google Scholar 

  8. Igney, F. & Krammer, P. Death and anti-death: tumour resistance to apoptosis. Nat. Rev. Cancer 2, 277–288 (2002).

    Article  CAS  Google Scholar 

  9. Ashkenazi, A. et al. Safety and antitumor activity of recombinant soluble Apo2 ligand. J. Clin. Invest. 104, 155–162 (1999).

    Article  CAS  Google Scholar 

  10. Kelley, S. & Ashkenazi, A. Targeting death receptors in cancer with Apo2L/TRAIL. Curr. Opin. Pharmacol. 4, 333–339 (2004).

    Article  CAS  Google Scholar 

  11. LeBlanc, H. et al. Tumor-cell resistance to death receptor–induced apoptosis through mutational inactivation of the proapoptotic Bcl-2 homolog Bax. Nat. Med. 8, 274–281 (2002).

    Article  CAS  Google Scholar 

  12. Hang, H. & Bertozzi, C. The chemistry and biology of mucin-type O-linked glycosylation. Bioorg. Med. Chem. 13, 5021–5034 (2005).

    Article  CAS  Google Scholar 

  13. Hanisch, F. O-glycosylation of the mucin type. Biol. Chem. 382, 143–149 (2001).

    Article  CAS  Google Scholar 

  14. Ohtsubo, K. & Marth, J.D. Glycosylation in cellular mechanisms of health and disease. Cell 126, 855–867 (2006).

    Article  CAS  Google Scholar 

  15. Ten Hagen, K.G., Fritz, T.A. & Tabak, L.A. All in the family: the UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferases. Glycobiology 13, 1R–16R (2003).

    Article  CAS  Google Scholar 

  16. Wang, H. et al. Cloning and characterization of a novel UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase, pp-GalNAc-T14. Biochem. Biophys. Res. Commun. 300, 738–744 (2003).

    Article  CAS  Google Scholar 

  17. Delannoy, P. et al. Benzyl-N-acetyl-α-D-galactosaminide inhibits the sialylation and the secretion of mucins by a mucin secreting HT-29 cell subpopulation. Glycoconj. J. 13, 717–726 (1996).

    Article  CAS  Google Scholar 

  18. Kuan, S.F., Byrd, J.C., Basbaum, C. & Kim, Y.S. Inhibition of mucin glycosylation by aryl-N-acetyl-α-galactosaminides in human colon cancer cells. J. Biol. Chem. 264, 19271–19277 (1989).

    CAS  PubMed  Google Scholar 

  19. Wei, M. et al. Proapoptotic BAX and BAK: a requisite gateway to mitochondrial dysfunction and death. Science 292, 727–730 (2001).

    Article  CAS  Google Scholar 

  20. Boldin, M.P. et al. Self-association of the “death domains” of the p55 tumor necrosis factor (TNF) receptor and Fas/APO1 prompts signaling for TNF and Fas/APO1 effects. J. Biol. Chem. 270, 387–391 (1995).

    Article  CAS  Google Scholar 

  21. Hansen, J.E. et al. NetOglyc: prediction of mucin type O-glycosylation sites based on sequence context and surface accessibility. Glycoconj. J. 15, 115–130 (1998).

    Article  CAS  Google Scholar 

  22. Hymowitz, S. et al. Triggering cell death: the crystal structure of Apo2L/TRAIL in a complex with death receptor 5. Mol. Cell 4, 563–571 (1999).

    Article  CAS  Google Scholar 

  23. Chan, F.K. et al. A domain in TNF receptors that mediates ligand-independent receptor assembly and signaling. Science 288, 2351–2354 (2000).

    Article  CAS  Google Scholar 

  24. Clancy, L. et al. Preligand assembly domain-mediated ligand-independent association between TRAIL receptor 4 (TR4) and TR2 regulates TRAIL-induced apoptosis. Proc. Natl. Acad. Sci. USA 102, 18099–18104 (2005).

    Article  CAS  Google Scholar 

  25. Feig, C., Tchikov, V., Schutze, S. & Peter, M.E. Palmitoylation of CD95 facilitates formation of SDS-stable receptor aggregates that initiate apoptosis signaling. EMBO J. 26, 221–231 (2007).

    Article  CAS  Google Scholar 

  26. Xu, Z. & Weiss, A. Negative regulation of CD45 by differential homodimerization of the alternatively spliced isoforms. Nat. Immunol. 3, 764–771 (2002).

    Article  CAS  Google Scholar 

  27. Haines, N. & Irvine, K.D. Glycosylation regulates Notch signalling. Nat. Rev. Mol. Cell Biol. 4, 786–797 (2003).

    Article  CAS  Google Scholar 

  28. Priatel, J.J. et al. The ST3Gal-I sialyltransferase controls CD8+ T lymphocyte homeostasis by modulating O-glycan biosynthesis. Immunity 12, 273–283 (2000).

    Article  CAS  Google Scholar 

  29. Brockhausen, I. Pathways of O-glycan biosynthesis in cancer cells. Biochim. Biophys. Acta 1473, 67–95 (1999).

    Article  CAS  Google Scholar 

  30. Dube, D. & Bertozzi, C. Glycans in cancer and inflammation—potential for therapeutics and diagnostics. Nat. Rev. Drug Discov. 4, 477–488 (2005).

    Article  CAS  Google Scholar 

  31. Fuster, M., Brown, J., Wang, L. & Esko, J. A disaccharide precursor of sialyl Lewis X inhibits metastatic potential of tumor cells. Cancer Res. 63, 2775–2781 (2003).

    CAS  PubMed  Google Scholar 

  32. Fuster, M. & Esko, J. The sweet and sour of cancer: glycans as novel therapeutic targets. Nat. Rev. Cancer 5, 526–542 (2005).

    Article  CAS  Google Scholar 

  33. Ohyama, C., Tsuboi, S. & Fukuda, M. Dual roles of sialyl Lewis X oligosaccharides in tumor metastasis and rejection by natural killer cells. EMBO J. 18, 1516–1525 (1999).

    Article  CAS  Google Scholar 

  34. Takada, A. et al. Contribution of carbohydrate antigens sialyl Lewis A and sialyl Lewis X to adhesion of human cancer cells to vascular endothelium. Cancer Res. 53, 354–361 (1993).

    CAS  PubMed  Google Scholar 

  35. Hoffman, E., Awad, T. & Palma, J. Expression profiling—best practices for data generation and interpretation in clinical trials. Nat. Rev. Genet. 5, 229–237 (2004).

    Article  CAS  Google Scholar 

  36. Yauch, R. et al. Epithelial versus mesenchymal phenotype determines in vitro sensitivity and predicts clinical activity of erlotinib in lung cancer patients. Clin. Cancer Res. 11, 8686–8698 (2005).

    Article  CAS  Google Scholar 

  37. Sharp, D.A., Lawrence, D.A. & Ashkenazi, A. Selective knockdown of the long variant of cellular FLICE inhibitory protein augments death receptor-mediated caspase-8 activation and apoptosis. J. Biol. Chem. 280, 19401–19409 (2005).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank S. Marsters and M. Nagel for plasmids and purification of recombinant DR5 respectively; S. Ross and M. Go for execution of xenograft studies; and W. Forrest for statistical analysis.

Author information

Authors and Affiliations

Authors

Contributions

K.W.W., D.L. and M.V.G. performed the cell line characterization. K.W.W. and G.C. performed the microarray analysis. E.A.P., T.J., D.A.L., R.M.P. and K.T. performed the functional and mechanistic studies. E.A.P., L.H., K.L. and V.K. performed the glycosylation and mass spectrometric analyses. S.F.Y. conducted the in vivo experiments. S.G.H. carried out the structural modeling. K.W.W., E.A.P., L.A. and A.A. guided the project and contributed to the experimental design and to data interpretation. K.W.W., E.A.P. and A.A. wrote the manuscript.

Corresponding author

Correspondence to Avi Ashkenazi.

Ethics declarations

Competing interests

All authors except K.W.W. and D.L. are presently employed by Genentech and may own Genentech shares.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–7, Supplementary Tables 1–2, Supplementary Methods (PDF 3522 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Wagner, K., Punnoose, E., Januario, T. et al. Death-receptor O-glycosylation controls tumor-cell sensitivity to the proapoptotic ligand Apo2L/TRAIL. Nat Med 13, 1070–1077 (2007). https://doi.org/10.1038/nm1627

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nm1627

This article is cited by

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