Interferon-γ increases the expression of glycosylated CD95 in B-leukemic cells

An inducible model to study the role of glycosylation in CD95-signalling and trafficking

Jan Dörrie, Katrin Sapala, Susan J. Zunino

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

B-lineage acute leukaemia cells are generally resistant to CD95-mediated apoptosis. In this report, the CD95-resistant B-leukaemia lines SEM, RS4;11, and REH were used to investigate the mechanisms of resistance to CD95-signalling. We found that interferon-γ (IFN-γ) treatment increased the presence of high molecular weight forms of CD95 in these cells as judged by Western analysis, and treatment of protein extracts with Peptide:N-glycosidase F indicated that the majority of high molecular weight forms were due to N-linked glycosylation. Treatment of whole cells with neuraminidase from Vibrio cholerae substantially reduced the relative molecular mass of CD95 observed after IFN-γ treatment and partially sensitized the three leukaemia lines to CD95-mediated death. To further characterize the different steps of oligosaccharide processing that may regulate CD95 signalling, the leukaemic cells were treated with IFN-γ and the glycosidase inhibitors castanospermine, 1-deoxymannojirimycin (DMM), and swainsonine. Treatment with DMM, a mannosidase inhibitor, efficiently reduced the appearance of high molecular weight forms of CD95 after IFN-γ treatment, and sensitized SEM and REH cells to CD95-mediated death. However, the IFN-γ-induced increases of CD95 on the cell surface were not altered by treatment with any of the glycosidase inhibitors, suggesting that the generation of complex oligosaccharide structures is not required for trafficking of CD95, but may instead be used as a mechanism of partially blocking CD95 signalling in these cells. In conclusion, IFN-γ treatment of the B-lineage leukaemia lines provides a novel, inducible system for the further characterization of post-translational modifications involved in modulating sensitivity to CD95-signalling.

Original languageEnglish (US)
Pages (from-to)98-107
Number of pages10
JournalCytokine
Volume18
Issue number2
DOIs
StatePublished - 2002
Externally publishedYes

Fingerprint

Glycosylation
Interferons
1-Deoxynojirimycin
Leukemia
Glycoside Hydrolases
Molecular weight
Oligosaccharides
Molecular Weight
Swainsonine
Mannosidases
Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase
Scanning electron microscopy
Neuraminidase
Molecular mass
Vibrio cholerae
Post Translational Protein Processing
Cells
Apoptosis
Processing
Proteins

Keywords

  • Apoptosis
  • B-lineage leukemia
  • CD95
  • Glycosylation
  • Interferon

ASJC Scopus subject areas

  • Endocrinology
  • Molecular Biology
  • Immunology
  • Immunology and Allergy

Cite this

Interferon-γ increases the expression of glycosylated CD95 in B-leukemic cells : An inducible model to study the role of glycosylation in CD95-signalling and trafficking. / Dörrie, Jan; Sapala, Katrin; Zunino, Susan J.

In: Cytokine, Vol. 18, No. 2, 2002, p. 98-107.

Research output: Contribution to journalArticle

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abstract = "B-lineage acute leukaemia cells are generally resistant to CD95-mediated apoptosis. In this report, the CD95-resistant B-leukaemia lines SEM, RS4;11, and REH were used to investigate the mechanisms of resistance to CD95-signalling. We found that interferon-γ (IFN-γ) treatment increased the presence of high molecular weight forms of CD95 in these cells as judged by Western analysis, and treatment of protein extracts with Peptide:N-glycosidase F indicated that the majority of high molecular weight forms were due to N-linked glycosylation. Treatment of whole cells with neuraminidase from Vibrio cholerae substantially reduced the relative molecular mass of CD95 observed after IFN-γ treatment and partially sensitized the three leukaemia lines to CD95-mediated death. To further characterize the different steps of oligosaccharide processing that may regulate CD95 signalling, the leukaemic cells were treated with IFN-γ and the glycosidase inhibitors castanospermine, 1-deoxymannojirimycin (DMM), and swainsonine. Treatment with DMM, a mannosidase inhibitor, efficiently reduced the appearance of high molecular weight forms of CD95 after IFN-γ treatment, and sensitized SEM and REH cells to CD95-mediated death. However, the IFN-γ-induced increases of CD95 on the cell surface were not altered by treatment with any of the glycosidase inhibitors, suggesting that the generation of complex oligosaccharide structures is not required for trafficking of CD95, but may instead be used as a mechanism of partially blocking CD95 signalling in these cells. In conclusion, IFN-γ treatment of the B-lineage leukaemia lines provides a novel, inducible system for the further characterization of post-translational modifications involved in modulating sensitivity to CD95-signalling.",
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