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Aberrant Glycosylation Augments the Immuno-Stimulatory Activities of Soluble Calreticulin

Calreticulin (CRT), a luminal resident calcium-binding glycoprotein of the cell, is a tumor-associated antigen involved in tumorigenesis and also an autoantigen targeted by autoantibodies found in patients with various autoimmune diseases. We have previously shown that prokaryotically expressed reco...

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Autores principales: Gong, Fang-Yuan, Gong, Zheng, Duo, Cui-Cui, Wang, Jun, Hong, Chao, Gao, Xiao-Ming
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6017544/
https://www.ncbi.nlm.nih.gov/pubmed/29495436
http://dx.doi.org/10.3390/molecules23030523
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author Gong, Fang-Yuan
Gong, Zheng
Duo, Cui-Cui
Wang, Jun
Hong, Chao
Gao, Xiao-Ming
author_facet Gong, Fang-Yuan
Gong, Zheng
Duo, Cui-Cui
Wang, Jun
Hong, Chao
Gao, Xiao-Ming
author_sort Gong, Fang-Yuan
collection PubMed
description Calreticulin (CRT), a luminal resident calcium-binding glycoprotein of the cell, is a tumor-associated antigen involved in tumorigenesis and also an autoantigen targeted by autoantibodies found in patients with various autoimmune diseases. We have previously shown that prokaryotically expressed recombinant murine CRT (rCRT) exhibits strong stimulatory activities against monocytes/macrophages in vitro and potent immunogenicity in vivo, which is partially attributable to self-oligomerization of soluble rCRT. However, even in oligomerized form native CRT (nCRT) isolated from mouse liver is much less active than rCRT, arguing against the possibility that self-oligomerization alone would license potent pro-inflammatory properties to nCRT. Since rCRT differs from nCRT in its lack of glycosylation, we wondered if aberrant glycosylation of eukaryotically expressed CRT (eCRT) would significantly enhance its immunological activity. In the present study, tunicamycin, an N-glycosyltransferase inhibitor, was employed to treat CHO cells (CHO-CRT) stably expressing full-length recombinant mouse CRT in secreted form for preparation of aberrantly glycosylated eCRT (tun-eCRT). Our biochemical and immunological analysis results indicate that eCRT produced by CHO-CRT cells is similar to nCRT in terms of glycosylation level, lack of self-oligomerization, relatively poor immunogenicity and weak macrophage-stimulatory activity, while tun-eCRT shows reduced glycosylation yet much enhanced ability to elicit specific humoral responses in mice and TNF-α and nitric oxide production by macrophages in vitro. Given that abberant glycosylation of proteins is a hallmark of cancer cells and also related to the development of autoimmune disorders in humans, our data may provide useful clues for better understanding of potentiating roles of dysregulated glycosylation of molecules such as CRT in tumorigenesis and autoimmunity.
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spelling pubmed-60175442018-11-13 Aberrant Glycosylation Augments the Immuno-Stimulatory Activities of Soluble Calreticulin Gong, Fang-Yuan Gong, Zheng Duo, Cui-Cui Wang, Jun Hong, Chao Gao, Xiao-Ming Molecules Article Calreticulin (CRT), a luminal resident calcium-binding glycoprotein of the cell, is a tumor-associated antigen involved in tumorigenesis and also an autoantigen targeted by autoantibodies found in patients with various autoimmune diseases. We have previously shown that prokaryotically expressed recombinant murine CRT (rCRT) exhibits strong stimulatory activities against monocytes/macrophages in vitro and potent immunogenicity in vivo, which is partially attributable to self-oligomerization of soluble rCRT. However, even in oligomerized form native CRT (nCRT) isolated from mouse liver is much less active than rCRT, arguing against the possibility that self-oligomerization alone would license potent pro-inflammatory properties to nCRT. Since rCRT differs from nCRT in its lack of glycosylation, we wondered if aberrant glycosylation of eukaryotically expressed CRT (eCRT) would significantly enhance its immunological activity. In the present study, tunicamycin, an N-glycosyltransferase inhibitor, was employed to treat CHO cells (CHO-CRT) stably expressing full-length recombinant mouse CRT in secreted form for preparation of aberrantly glycosylated eCRT (tun-eCRT). Our biochemical and immunological analysis results indicate that eCRT produced by CHO-CRT cells is similar to nCRT in terms of glycosylation level, lack of self-oligomerization, relatively poor immunogenicity and weak macrophage-stimulatory activity, while tun-eCRT shows reduced glycosylation yet much enhanced ability to elicit specific humoral responses in mice and TNF-α and nitric oxide production by macrophages in vitro. Given that abberant glycosylation of proteins is a hallmark of cancer cells and also related to the development of autoimmune disorders in humans, our data may provide useful clues for better understanding of potentiating roles of dysregulated glycosylation of molecules such as CRT in tumorigenesis and autoimmunity. MDPI 2018-02-27 /pmc/articles/PMC6017544/ /pubmed/29495436 http://dx.doi.org/10.3390/molecules23030523 Text en © 2018 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Gong, Fang-Yuan
Gong, Zheng
Duo, Cui-Cui
Wang, Jun
Hong, Chao
Gao, Xiao-Ming
Aberrant Glycosylation Augments the Immuno-Stimulatory Activities of Soluble Calreticulin
title Aberrant Glycosylation Augments the Immuno-Stimulatory Activities of Soluble Calreticulin
title_full Aberrant Glycosylation Augments the Immuno-Stimulatory Activities of Soluble Calreticulin
title_fullStr Aberrant Glycosylation Augments the Immuno-Stimulatory Activities of Soluble Calreticulin
title_full_unstemmed Aberrant Glycosylation Augments the Immuno-Stimulatory Activities of Soluble Calreticulin
title_short Aberrant Glycosylation Augments the Immuno-Stimulatory Activities of Soluble Calreticulin
title_sort aberrant glycosylation augments the immuno-stimulatory activities of soluble calreticulin
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6017544/
https://www.ncbi.nlm.nih.gov/pubmed/29495436
http://dx.doi.org/10.3390/molecules23030523
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