Differential profiling studies of N-linked glycoproteins in glioblastoma cancer stem cells upon treatment with γ-secretase inhibitor

Program of Bioinformatics, University of Michigan Medical Center, Ann Arbor, MI 48109-0650, USA.
Proteomics (Impact Factor: 3.81). 10/2011; 11(20):4021-8. DOI: 10.1002/pmic.201100014
Source: PubMed


We have recently demonstrated that Notch pathway blockade by γ-secretase inhibitor (GSI) depletes cancer stem cells (CSCs) in Glioblastoma Multiforme (GBM) through reduced proliferation and induced apoptosis. However, the detailed mechanism by which the manipulation of Notch signal induces alterations on post-translational modifications such as glycosylation has not been investigated. Herein, we present a differential profiling work to detect the change of glycosylation pattern upon drug treatment in GBM CSCs. Rapid screening of differential cell surface glycan structures has been performed by lectin microarray on live cells followed by the detection of N-linked glycoproteins from cell lysates using multi-lectin chromatography and label-free quantitative mass spectrometry analysis. A total of 51 and 52 glycoproteins were identified in the CSC- and GSI-treated groups, respectively, filtered by a combination of decoy database searching and Trans-Proteomic Pipeline (TPP) processing. Although no significant changes were detected from the lectin microarray experiment, 7 differentially expressed glycoproteins with high confidence were captured after the multi-lectin column including key enzymes involved in glycan processing. Functional annotations of the altered glycoproteins suggest a phenotype transformation of CSCs toward a less tumorigenic form upon GSI treatment.

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    • "d for identification of serum biomarkers for lung adenocarcinoma ( Heo et al . , 2007 ) , breast cancer ( Zeng et al . , 2010b ) , and ovarian cancer ( Abbott et al . , 2010 ) . Differential label - free profiling studies of N - linked glycoproteins in cancer stem cells were tried to detect the change of glycosylation pattern upon drug treatment ( Dai et al . , 2011 ; He et al . , 2011 ) ."
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    ABSTRACT: Mass spectrometry (MS) has been a core technology for high sensitive and high-throughput analysis of the enriched glycoproteome in aspects of quantitative assays as well as qualitative profiling of glycoproteins. Because it has been widely recognized that aberrant glycosylation in a glycoprotein may involve in progression of a certain disease, the development of efficient analysis tool for the aberrant glycoproteins is very important for deep understanding about pathological function of the glycoprotein and new biomarker development. This review first describes the protein glycosylation-targeting enrichment technologies mainly employing solid-phase extraction methods such as hydrizide-capturing, lectin-specific capturing, and affinity separation techniques based on porous graphitized carbon, hydrophilic interaction chromatography, or immobilized boronic acid. Second, MS-based quantitative analysis strategies coupled with the protein glycosylation-targeting enrichment technologies, by using a label-free MS, stable isotope-labeling, or targeted multiple reaction monitoring (MRM) MS, are summarized with recent published studies. © 2014 The Authors. Mass Spectrometry Reviews Published by Wiley Periodicals, Inc. Rapid Commun. Mass Spectrom.
    Mass Spectrometry Reviews 04/2015; 34(2). DOI:10.1002/mas.21428 · 7.71 Impact Factor
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    • "Unfortunately, the information of binding sites of carbohydrates on proteins could be lost. The other approach is to directly digest glycoproteins with endoprotease, and then digested glycopeptides are characterized by mass spectrometry [28]. The glycosylation sites can be determined. "
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    ABSTRACT: Glycosylation is one of the most important posttranslational modifications of proteins and plays essential roles in various biological processes. Aberration in the glycan moieties of glycoproteins is associated with many diseases. It is especially critical to develop the rapid and sensitive methods for analysis of aberrant glycoproteins associated with diseases. Mass spectrometry (MS) has become a powerful tool for glycoprotein analysis. Especially, tandem mass spectrometry can provide highly informative fragments for structural identification of glycoproteins. This review provides an overview of the development of MS technologies and their applications in identification of abnormal glycoproteins and glycans in human serum to screen cancer biomarkers in recent years.
    Clinical Proteomics 04/2014; 11(1):14. DOI:10.1186/1559-0275-11-14
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    ABSTRACT: Normal multipotent tissue stem cells (SCs) are the driving force behind tissue turnover and repair. The cancer stem cell theory holds that tumors also contain stem-like cells that drive tumor growth and metastasis formation. However, very little is known about the regulation of SC maintenance pathways in cancer and how these are affected by cancer-specific genetic alterations and by treatment. Proteomics is emerging as a powerful tool to identify the signaling complexes and pathways that control multi- and pluri-potency and SC differentiation. Here, the authors review the novel insights that these studies have provided and present a comprehensive strategy for the use of proteomics in studying cancer SC biology.
    Expert Review of Proteomics 06/2012; 9(3):325-36. DOI:10.1586/epr.12.24 · 2.90 Impact Factor
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