Altered glycosylation of recombinant NKp30 hampers binding to heparan sulfate: a lesson for the use of recombinant immunoreceptors as an immunological tool.

2.50
Hdl Handle:
http://hdl.handle.net/10541/56020
Title:
Altered glycosylation of recombinant NKp30 hampers binding to heparan sulfate: a lesson for the use of recombinant immunoreceptors as an immunological tool.
Authors:
Hershkovitz, Oren; Jarahian, Mostafa; Zilka, Alon; Bar-Ilan, Ahuva; Landau, Guy; Jivov, Sergey; Tekoah, Yoram; Glicklis, Rachel; Gallagher, John T; Hoffmann, Sabrina C; Zer, Hagit; Mandelboim, Ofer; Watzl, Carsten; Momburg, Frank; Porgador, Angel
Abstract:
NKp30 is a natural cytotoxicity receptor expressed by human NK cells and involved in NK lytic activity. We previously published that membranal heparan sulfate serves as a coligand for human NKp30. In the present study, we complement our results by showing direct binding of recombinant NKp30 to immobilized heparin. The heparan sulfate epitope(s) on target tumor cells and the heparin epitope(s) recognized by NKp30 share similar characteristics. Warren and colleagues (Warren HS, Jones AL, Freeman C, Bettadapura J, Parish CR. 2005. Evidence that the cellular ligand for the human NK cell activation receptor NKp30 is not a heparan sulfate glycosaminoglycan. J Immunol. 175:207-212) published that NKp30 does not bind to membranal heparan sulfate on target cells and that heparan sulfate is not involved in NKp30-mediated lysis. In the current study, we examine the binding of six different recombinant NKp30s to membranal heparan sulfate and conclude that NKp30 does interact with membranal heparan sulfate. Yet, two of the six recombinant NKp30s, including the commercially available recombinant NKp30 (employed by Warren et al.) did not show heparan sulfate-dependent binding. We demonstrate that this is due to an altered glycosylation of these two recombinant NKp30s. Upon removal of its N-linked glycans, heparan sulfate-dependent binding to tumor cells and direct binding to heparin were restored. Overall, our results emphasize the importance of proper glycosylation for analysis of NKp30 binding to its ligand and that membranal heparan sulfate could serve as a coligand for NKp30. At the cellular level, soluble heparan sulfate enhanced the secretion of IFNgamma by NK-92 natural killer cells activated with anti-NKp30 monoclonal antibody. We discuss the involvement of heparan sulfate binding to NKp30 in NKp30-mediated activation of NK cells.
Affiliation:
The Shraga Segal Department of Microbiology and Immunology, Faculty of Health Sciences and the Cancer Research Center, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel.
Citation:
Altered glycosylation of recombinant NKp30 hampers binding to heparan sulfate: a lesson for the use of recombinant immunoreceptors as an immunological tool. 2008, 18 (1):28-41 Glycobiology
Journal:
Glycobiology
Issue Date:
Jan-2008
URI:
http://hdl.handle.net/10541/56020
DOI:
10.1093/glycob/cwm125
PubMed ID:
18006589
Type:
Article
Language:
en
ISSN:
1460-2423
Appears in Collections:
All Paterson Institute for Cancer Research; Medical Oncology

Full metadata record

DC FieldValue Language
dc.contributor.authorHershkovitz, Oren-
dc.contributor.authorJarahian, Mostafa-
dc.contributor.authorZilka, Alon-
dc.contributor.authorBar-Ilan, Ahuva-
dc.contributor.authorLandau, Guy-
dc.contributor.authorJivov, Sergey-
dc.contributor.authorTekoah, Yoram-
dc.contributor.authorGlicklis, Rachel-
dc.contributor.authorGallagher, John T-
dc.contributor.authorHoffmann, Sabrina C-
dc.contributor.authorZer, Hagit-
dc.contributor.authorMandelboim, Ofer-
dc.contributor.authorWatzl, Carsten-
dc.contributor.authorMomburg, Frank-
dc.contributor.authorPorgador, Angel-
dc.date.accessioned2009-03-17T17:05:43Z-
dc.date.available2009-03-17T17:05:43Z-
dc.date.issued2008-01-
dc.identifier.citationAltered glycosylation of recombinant NKp30 hampers binding to heparan sulfate: a lesson for the use of recombinant immunoreceptors as an immunological tool. 2008, 18 (1):28-41 Glycobiologyen
dc.identifier.issn1460-2423-
dc.identifier.pmid18006589-
dc.identifier.doi10.1093/glycob/cwm125-
dc.identifier.urihttp://hdl.handle.net/10541/56020-
dc.description.abstractNKp30 is a natural cytotoxicity receptor expressed by human NK cells and involved in NK lytic activity. We previously published that membranal heparan sulfate serves as a coligand for human NKp30. In the present study, we complement our results by showing direct binding of recombinant NKp30 to immobilized heparin. The heparan sulfate epitope(s) on target tumor cells and the heparin epitope(s) recognized by NKp30 share similar characteristics. Warren and colleagues (Warren HS, Jones AL, Freeman C, Bettadapura J, Parish CR. 2005. Evidence that the cellular ligand for the human NK cell activation receptor NKp30 is not a heparan sulfate glycosaminoglycan. J Immunol. 175:207-212) published that NKp30 does not bind to membranal heparan sulfate on target cells and that heparan sulfate is not involved in NKp30-mediated lysis. In the current study, we examine the binding of six different recombinant NKp30s to membranal heparan sulfate and conclude that NKp30 does interact with membranal heparan sulfate. Yet, two of the six recombinant NKp30s, including the commercially available recombinant NKp30 (employed by Warren et al.) did not show heparan sulfate-dependent binding. We demonstrate that this is due to an altered glycosylation of these two recombinant NKp30s. Upon removal of its N-linked glycans, heparan sulfate-dependent binding to tumor cells and direct binding to heparin were restored. Overall, our results emphasize the importance of proper glycosylation for analysis of NKp30 binding to its ligand and that membranal heparan sulfate could serve as a coligand for NKp30. At the cellular level, soluble heparan sulfate enhanced the secretion of IFNgamma by NK-92 natural killer cells activated with anti-NKp30 monoclonal antibody. We discuss the involvement of heparan sulfate binding to NKp30 in NKp30-mediated activation of NK cells.en
dc.language.isoenen
dc.subjectGlycosylationen
dc.subjectHeparan Sulfateen
dc.subjectNatural Killeren
dc.subjectNCRen
dc.subjectNKp30en
dc.subjectCell Line, Tumour-
dc.subject.meshAnimals-
dc.subject.meshBinding Sites-
dc.subject.meshCHO Cells-
dc.subject.meshCell Line, Tumor-
dc.subject.meshCricetinae-
dc.subject.meshCricetulus-
dc.subject.meshGlycosylation-
dc.subject.meshHela Cells-
dc.subject.meshHeparitin Sulfate-
dc.subject.meshHumans-
dc.subject.meshNatural Cytotoxicity Triggering Receptor 3-
dc.subject.meshPeptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase-
dc.subject.meshPolysaccharides-
dc.subject.meshReceptors, Immunologic-
dc.subject.meshRecombinant Proteins-
dc.subject.meshSpectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization-
dc.titleAltered glycosylation of recombinant NKp30 hampers binding to heparan sulfate: a lesson for the use of recombinant immunoreceptors as an immunological tool.en
dc.typeArticleen
dc.contributor.departmentThe Shraga Segal Department of Microbiology and Immunology, Faculty of Health Sciences and the Cancer Research Center, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel.en
dc.identifier.journalGlycobiologyen

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