Glycans expand the structural complexity of proteins by several orders of magnitude, resulting in a tremendous analytical challenge when including them in biomedical research. Recent... Show moreGlycans expand the structural complexity of proteins by several orders of magnitude, resulting in a tremendous analytical challenge when including them in biomedical research. Recent glycobiological research is painting a picture in which glycans represent a crucial structural and functional component of the majority of proteins, with alternative glycosylation of proteins and lipids being an important regulatory mechanism in many biological and pathological processes. Since interindividual differences in glycosylation are extensive, large studies are needed to map the structures and to understand the role of glycosylation in human (patho)physiology. Driven by these challenges, methods have emerged, which can tackle the complexity of glycosylation in thousands of samples, also known as high-throughput (HT) glycomics. For facile dissemination and implementation of HT glycomics technology, the sample preparation, analysis, as well as data mining, need to be stable over a long period of time (months/years), amenable to automation, and available to non-specialized laboratories. Current HT glycomics methods mainly focus on protein N-glycosylation and allow to extensively characterize this subset of the human glycome in large numbers of various biological samples. The ultimate goal in HT glycomics is to gain better knowledge and understanding of the complete human glycome using methods that are easy to adapt and implement in (basic) biomedical research. Aiming to promote wider use and development of HT glycomics, here, we present currently available, emerging, and prospective methods and some of their applications, revealing a largely unexplored molecular layer of the complexity of life. Show less
Aberrant expression of certain glycosphingolipids (GSLs) is associated withthe differentiation of acute myeloid leukemia (AML) cells. However, the expressionpatterns of GSLs in AML are still poorly... Show moreAberrant expression of certain glycosphingolipids (GSLs) is associated withthe differentiation of acute myeloid leukemia (AML) cells. However, the expressionpatterns of GSLs in AML are still poorly explored because of their complexity, the presenceof multiple isomeric structures, and tedious analytical procedures. In this study, weperformed an in-depth GSL glycan analysis of 19 AML cell lines using porous graphitizedcarbon liquid chromatography-mass spectrometry revealing strikingly different GSL glycanprofiles between the various AML cell lines. The cell lines of the M6 subtype showed a highexpression of gangliosides with alpha 2,3-sialylation and Neu5Gc, while the M2 and M5subtypes were characterized by high expression of (neo)lacto-series glycans and Lewis A/Xantigens. Integrated analysis of glycomics and available transcriptomics data revealed theassociation of GSL glycan abundances with the transcriptomics expression of certainglycosyltransferases (GTs) and transcription factors (TFs). In addition, correlations werefound between specific GTs and TFs. Our data reveal TFsGATA2,GATA1, andRUNX1as candidate inducers of the expression of gangliosides and sialylation via regulation of the GTsST3GAL2andST8SIA1.Inconclusion, we show that GSL glycan expression levels are associated with hematopoietic AML classifications and TF and GT geneexpression. Further research is needed to dissect the regulation of GSL expression and its role in hematopoiesis and associated malignancies. Show less
Background: Immunoglobulin G1 (IgG1) effector functions are impacted by the structure of fragment crystallizable (Fc) tail-linked N-glycans. Low fucosylation levels on severe acute respiratory... Show moreBackground: Immunoglobulin G1 (IgG1) effector functions are impacted by the structure of fragment crystallizable (Fc) tail-linked N-glycans. Low fucosylation levels on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) protein-specific IgG1 has been described as a hallmark of severe coronavirus disease 2019 (COVID-19) and may lead to activation of macrophages via immune complexes thereby promoting inflammatory responses, altogether suggesting involvement of IgG1 Fc glycosylation modulated immune mechanisms in COVID-19. Methods: In this prospective, observational single center cohort study, IgG1 Fc glycosylation was analyzed by liquid chromatography-mass spectrometry following affinity capturing from serial plasma samples of 159 SARS-CoV-2 infected hospitalized patients. Findings: At baseline close to disease onset, anti-S IgG1 glycosylation was highly skewed when compared to total plasma IgG1. A rapid, general reduction in glycosylation skewing was observed during the disease course. Low anti S IgG1 galactosylation and sialylation as well as high bisection were early hallmarks of disease severity, whilst high galactosylation and sialylation and low bisection were found in patients with low disease severity. In line with these observations, anti-S IgG1 glycosylation correlated with various inflammatory markers. Interpretation: Association of low galactosylation, sialylation as well as high bisection with disease severity and inflammatory markers suggests that further studies are needed to understand how anti-S IgG1 glycosylation may contribute to disease mechanism and to evaluate its biomarker potential. Copyright (c) 2022 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/) Show less
Nicolardi, S.; Danuser, R.; Dotz, V.; Dominguez Vega, E.; Kaabi, A. al; Beurret, M.; ... ; Wuhrer, M. 2022
Bacterial glycoconjugate vaccines have a major role in preventing microbial infections. Immunogenic bacterial glycans, suchas O-antigen polysaccharides, can be recombinantly expressed and combined... Show moreBacterial glycoconjugate vaccines have a major role in preventing microbial infections. Immunogenic bacterial glycans, suchas O-antigen polysaccharides, can be recombinantly expressed and combined with specific carrier proteins to produce effective vaccines. O-Antigen polysaccharides are typically polydisperse, and carrier proteins can have multiple glycosylation sites. Consequently, recombinant glycoconjugate vaccines have a high structural heterogeneity, making their characterization challenging. Sincedevelopment and quality control processes rely on such character-ization, novel strategies are needed for faster and informative analysis.Here, we present a novel approach employing minimal samplepreparation and ultrahigh-resolution mass spectrometry analysis forprotein terminal sequencing and characterization of the oligosaccharide repeat units of bacterial glycoconjugate vaccines. Threeglycoconjugate vaccine candidates, obtained from the bioconjugation of the O-antigen polysaccharides fromE. coliserotypes O2,O6A, and O25B with the genetically detoxified exotoxin A fromPseudomonas aeruginosa, were analyzed by MALDI-in-source decay(ISD) FT-ICR MS. Protein and glycan ISD fragment ions were selectively detected using 1,5-diaminonaphtalene and a 2,5-dihydroxybenzoic acid/2-hydroxy-5-methoxybenzoic acid mixture (super-DHB) as a MALDI matrix, respectively. The analysis of protein fragments required the absence of salts in the samples, while the presence of salt was key for the detection of sodiated glycanfragments. MS/MS analysis of O-antigen ISD fragments allowed for the detection of specific repeat unit signatures. The developed strategy requires minute sample amounts, avoids the use of chemical derivatizations, and comes with minimal hands-on time allowing for fast corroboration of key structural features of bacterial glycoconjugate vaccines during early- and late-stage development Show less
Immune-mediated platelet refractoriness (PR) remains a significant problem in the setting of platelet transfusion and is predominantly caused by the presence of alloantibodies directed against... Show moreImmune-mediated platelet refractoriness (PR) remains a significant problem in the setting of platelet transfusion and is predominantly caused by the presence of alloantibodies directed against class I human leukocyte antigens (HLA). Opsonization of donor platelets with these alloantibodies can result in rapid clearance after transfusion via multiple mechanisms, including antibody dependent cellular phagocytosis (ADCP). Interestingly, not all alloimmunized patients develop PR to unmatched platelet transfusions, suggesting variation in HLA-specific IgG responses between patients. Previously, we observed that the glycosylation profile of anti-HLA antibodies was highly variable between PR patients, especially with respect to Fc galactosylation, sialylation and fucosylation. In the current study, we investigated the effect of different Fc glycosylation patterns, with known effects on complement deposition and FcγR binding, on phagocytosis of opsonized platelets by monocyte-derived human macrophages. We found that the phagocytosis of antibody- and complement-opsonized platelets, by monocyte derived M1 macrophages, was unaffected by these qualitative IgG-glycan differences. Show less
Zhang, J.; Zhang, Z.J.; Holst, S.; Bloechl, C.; Madunic, K.; Wuhrer, M.; ... ; Zhang, T. 2022
Pancreatic ductal adenocarcinoma (PDAC) is characterized by poor prognosis and high mortality. Transforming growth factor-beta (TGF-beta) plays a key role in PDAC tumor progression, which is often... Show morePancreatic ductal adenocarcinoma (PDAC) is characterized by poor prognosis and high mortality. Transforming growth factor-beta (TGF-beta) plays a key role in PDAC tumor progression, which is often associated with aberrant glycosylation. However, how PDAC cells respond to TGF-beta and the role of glycosylation therein is not well known. Here, we investigated the TGF-beta-mediated response and glycosylation changes in the PaTu-8955S (PaTu-S) cell line deficient in SMA-related and MAD-related protein 4 (SMAD4), a signal transducer of the TGF-beta signaling. PaTu-S cells responded to TGF-beta by upregulating SMAD2 phosphorylation and target gene expression. We found that TGF-beta induced expression of the mesenchymal marker N-cadherin but did not significantly affect epithelial marker E-cadherin expression. We also examined differences in N-glycans, O-glycans, and glycosphingolipid-linked glycans in PaTu-S cells upon TGF-beta stimulation. TGF-beta treatment primarily induced N-glycome aberrations involving elevated levels of branching, core fucosylation, and sialylation in PaTu-S cells, in agreement with TGF-beta-induced changes in the expression of glycosylation-associated genes. In addition, we observed differences in O glycosylation and glycosphingolipid glycosylation profiles after TGF-beta treatment, including lower levels of sialylated Tn antigen and neoexpression of globosides. Furthermore, the expression of transcription factor sex-determining region Y-related high-mobility group box 4 was upregulated upon TGF-beta stimulation, and its depletion blocked TGF-beta-induced N-glycomic changes. Thus, TGF-beta-induced N-glycosylation changes can occur in a sex-determining region Y-related high-mobility group box 4-dependent and SMAD4-independent manner in the pancreatic PaTu-S cancer cell line. Our results open up avenues to study the relevance of glycosylation in TGF-beta signaling in SMAD4-inactivated PDAC. Show less
Boyaval, F.; Dalebout, H.; Zeijl, R. van; Wang, W.J.; Farina-Sarasqueta, A.; Lageveen-Kammeijer, G.S.M.; ... ; Heijs, B. 2022
Simple Summary The detection of colorectal cancer (CRC) at an early stage is increasing due to the implementation of screening programs. Local excision of early CRC is potentially curative, however... Show moreSimple Summary The detection of colorectal cancer (CRC) at an early stage is increasing due to the implementation of screening programs. Local excision of early CRC is potentially curative, however the identification of early lesions at high risk of regional metastases remains challenging, and greatly influencing therapy decision making. Variations in sugar molecules has been associated with development and progression in various cancer types including CRC. Therefore, we examined these sugar signatures, so-called N-glycans, in different stages of progression of CRC starting from epithelium to pre-cancerous and cancerous tissue. We report that the sugar signatures clearly differentiate each step of CRC progression, especially between pre-cancerous and cancerous tissue. We also observed some of the glycosylation signatures of the cancerous areas to be spreading into the tumor microenvironment. The increase incidence of early colorectal cancer (T1 CRC) last years is mainly due to the introduction of population-based screening for CRC. T1 CRC staging based on histological criteria remains challenging and there is high variability among pathologists in the scoring of these criteria. It is crucial to unravel the biology behind the progression of adenoma into T1 CRC. Glycomic studies have reported extensively on alterations of the N-glycomic pattern in CRC; therefore, investigating these alterations may reveal new insights into the development of T1 CRC. We used matrix-assisted laser desorption ionization (MALDI) mass spectrometry imaging (MSI) to spatially profile the N-glycan species in a cohort of pT1 CRC using archival formalin-fixed and paraffin-embedded (FFPE) material. To generate structural information on the observed N-glycans, CE-ESI-MS/MS was used in conjunction with MALDI-MSI. Relative intensities and glycosylation traits were calculated based on a panel of 58 N-glycans. Our analysis showed pronounced differences between normal epithelium, dysplastic, and carcinoma regions. High-mannose-type N-glycans were higher in the dysplastic region than in carcinoma, which correlates to increased proliferation of the cells. We observed changes in the cancer invasive front, including higher expression of alpha 2,3-linked sialic acids which followed the glycosylation pattern of the carcinoma region. Show less
Human immunodeficiency virus 1 (HIV-1) virus-like particles (VLPs) are nanostructures derived from the self-assembly and cell budding of Gag polyprotein. Mimicking the native structure of the virus... Show moreHuman immunodeficiency virus 1 (HIV-1) virus-like particles (VLPs) are nanostructures derived from the self-assembly and cell budding of Gag polyprotein. Mimicking the native structure of the virus and being noninfectious, they represent promising candidates for the development of new vaccines as they elicit a strong immune response. In addition to this, the bounding membrane can be functionalized with exogenous antigens to target different diseases. Protein glycosylation depends strictly on the production platform and expression system used and the displayed glycosylation patterns may influence downstream processing as well as the immune response. One of the main challenges for the development of Gag VLP production bioprocess is the separation of VLPs and coproduced extracellular vesicles (EVs). In this study, porous graphitized carbon separation method coupled with mass spectrometry was used to characterize the N- and O- glycosylation profiles of Gag VLPs produced in HEK293 cells. We identified differential glycan signatures between VLPs and EVs that could pave the way for further separation and purification strategies to optimize downstream processing and move forward in VLP-based vaccine production technology. Show less
The hallmark autoantibodies in rheumatoid arthritis are characterized by variable domain glycans (VDGs). Their abundant occurrence results from the selective introduction of N-linked glycosylation... Show moreThe hallmark autoantibodies in rheumatoid arthritis are characterized by variable domain glycans (VDGs). Their abundant occurrence results from the selective introduction of N-linked glycosylation sites during somatic hypermutation, and their presence is predictive for disease development. However, the functional consequences of VDGs on autoreactive B cells remain elusive. Combining crystallography, glycobiology, and functional B cell assays allowed us to dissect key characteristics of VDGs on human B cell biology. Crystal structures showed that VDGs are positioned in the vicinity of the antigen-binding pocket, and dynamic modeling combined with binding assays elucidated their impact on binding. We found that VDG-expressing B cell receptors stay longer on the B cell surface and that VDGs enhance B cell activation. These results provide a rationale on how the acquisition of VDGs might contribute to the breach of tolerance of autoreactive B cells in a major human autoimmune disease. Show less
Lipopolysaccharides, the major outer membrane components of Gram-negative bacteria, are crucial actors of the host-microbial dialogue. They can contribute to the establishment of either symbiosis... Show moreLipopolysaccharides, the major outer membrane components of Gram-negative bacteria, are crucial actors of the host-microbial dialogue. They can contribute to the establishment of either symbiosis or bacterial virulence, depending on the bacterial lifestyle. Plant microbiota shows great complexity, promotes plant health and growth and assures protection from pathogens. How plants perceive LPS from plant-associated bacteria and discriminate between beneficial and pathogenic microbes is an open and urgent question. Here, we report on the structure, conformation, membrane properties and immune recognition of LPS isolated from the Arabidopsis thaliana root microbiota member Herbaspirillum sp. Root189. The LPS consists of an O-methylated and variously acetylated Drhamnose containing polysaccharide with a rather hydrophobic surface. Plant immunology studies in A. thaliana demonstrate that the native acetylated O-antigen shields the LPS from immune recognition whereas the O- deacylated one does not. These findings highlight the role of Herbaspirillum LPS within plant-microbial crosstalk, and how O-antigen modifications influence membrane properties and modulate LPS host recognition. Show less
The hallmark autoantibodies in rheumatoid arthritis are characterized by variable domain glycans (VDGs). Their abundant occurrence results from the selective introduction of N-linked glycosylation... Show moreThe hallmark autoantibodies in rheumatoid arthritis are characterized by variable domain glycans (VDGs). Their abundant occurrence results from the selective introduction of N-linked glycosylation sites during somatic hyper-mutation, and their presence is predictive for disease development. However, the functional consequences of VDGs on autoreactive B cells remain elusive. Combining crystallography, glycobiology, and functional B cell assays allowed us to dissect key characteristics of VDGs on human B cell biology. Crystal structures showed that VDGs are positioned in the vicinity of the antigen-binding pocket, and dynamic modeling combined with binding assays elucidated their impact on binding. We found that VDG-expressing B cell receptors stay longer on the B cell surface and that VDGs enhance B cell activation. These results provide a rationale on how the acquisition of VDGs might contribute to the breach of tolerance of autoreactive B cells in a major human autoimmune disease. Show less
Schaick, G. van; Hajjouti, N. el; Nicolardi, S.; Hartog, J. den; Jansen, R.; Hoeven, R. van der; ... ; Dominguez Vega, E. 2022
Xylanases are of great value in various industries, including paper, food, and biorefinery. Due to their biotechnological production, these enzymes can contain a variety of post-translational... Show moreXylanases are of great value in various industries, including paper, food, and biorefinery. Due to their biotechnological production, these enzymes can contain a variety of post-translational modifications, which may have a profound effect on protein function. Understanding the structure-function relationship can guide the development of products with optimal performance. We have developed a workflow for the structural and functional characterization of an endo-1,4-beta-xylanase (ENDO-I) produced by Aspergillus niger with and without applying thermal stress. This workflow relies on orthogonal native separation techniques to resolve proteoforms. Mass spectrometry and activity assays of separated proteoforms permitted the establishment of structure-function relationships. The separation conditions were focus on balancing efficient separation and protein functionality. We employed size exclusion chromatography (SEC) to separate ENDO-I from other co-expressed proteins. Charge variants were investigated with ion exchange chromatography (IEX) and revealed the presence of low abundant glycated variants in the temperature-stressed material. To obtain better insights into the effect on glycation on function, we enriched for these species using boronate affinity chromatography (BAC). The activity measurements showed lower activity of glycated species compared to the non-modified enzyme. Altogether, this workflow allowed in-depth structural and functional characterization of ENDO-I proteoforms. Show less
Schaick, G. van; Haselberg, R.; Somsen, G.W.; Wuhrer, M.; Dominguez Vega, E. 2022
Alterations in protein structure may have profound effects on biological function. Analytical techniques that permit characterization of proteins while maintaining their conformational and... Show moreAlterations in protein structure may have profound effects on biological function. Analytical techniques that permit characterization of proteins while maintaining their conformational and functional state are crucial for studying changes in the higher order structure of proteins and for establishing structure-function relationships. Coupling of native protein separations with mass spectrometry is emerging rapidly as a powerful approach to study these aspects in a reliable, fast and straightforward way. This Review presents the available native separation modes for proteins, covers practical considerations on the hyphenation of these separations with mass spectrometry and highlights the involvement of affinity-based separations to simultaneously obtain structural and functional information of proteins. The impact of these approaches is emphasized by selected applications addressing biomedical and biopharmaceutical research questions. Show less
Grunert, I.; Heinrich, K.; Ernst, J.; Hingar, M.; Briguet, A.; Leiss, M.; ... ; Bulau, P. 2022
In recent years, a variety of new antibody formats have been developed. One of these formats allows the binding of one type of antibody to two different epitopes. This can for example be achieved... Show moreIn recent years, a variety of new antibody formats have been developed. One of these formats allows the binding of one type of antibody to two different epitopes. This can for example be achieved by introduction of the "knob-into-hole" format and a combined CrossMab approach. Due to their complexity, these bispecific antibodies are expected to result in an enhanced variety of different degradation products. Reports on the stability of these molecules are still largely lacking. To address this, a panel of stress conditions, including elevated temperature, pH, oxidizing agents, and forced glycation via glucose incubation, to identify and functionally evaluate critical quality attributes in the complementary-determining and conserved regions of a bispecific antibody was applied in this study. The exertion of various stress conditions combined with an assessment by size exclusion chromatography, ion exchange chromatography, LC-MS/MS peptide mapping, and functional evaluation by cell-based assays was adequate to identify chemical modification sites and assess the stability and integrity, as well as the functionality of a bispecific antibody. Stress conditions induced size variants and post-translational modifications, such as isomerization, deamidation, and oxidation, albeit to a modest extent. Of note, all the observed stress conditions largely maintained functionality. In summary, this study revealed the pronounced stability of IgG1 "knob-into-hole" bispecific CrossMab antibodies compared to already marketed antibody products. Show less
Rodriguez, E.; Boelaars, K.; Brown, K.; Madunic, K.; Ee, T. van; Dijk, F.; ... ; Kooyk, Y. van 2022
Pancreatic ductal adenocarcinoma (PDAC) remains one of the most aggressive malignancies with a 5-year survival rate of only 9%. Despite the fact that changes in glycosylation patterns during tumour... Show morePancreatic ductal adenocarcinoma (PDAC) remains one of the most aggressive malignancies with a 5-year survival rate of only 9%. Despite the fact that changes in glycosylation patterns during tumour progression have been reported, no systematic approach has been conducted to evaluate its potential for patient stratification. By analysing publicly available transcriptomic data of patient samples and cell lines, we identified here two specific glycan profiles in PDAC that correlated with progression, clinical outcome and epithelial to mesenchymal transition (EMT) status. These different glycan profiles, confirmed by glycomics, can be distinguished by the expression of O-glycan fucosylated structures, present only in epithelial cells and regulated by the expression of GALNT3. Moreover, these fucosylated glycans can serve as ligands for DC-SIGN positive tumour-associated macrophages, modulating their activation and inducing the production of IL-10. Our results show mechanisms by which the glyco-code contributes to the tolerogenic microenvironment in PDAC.Rodriguez et al. present a transcriptomic analysis of glycosylation associated gene profiles, including bulk patient sequencing, sc-RNA seq, cell lines and organoids, to examine glycosylation in PDAC. They find 2 specific glycan profiles correlating with progression, clinical outcome and EMT, and conclude that the glyco-code contributes to the tolerogenic microenvironment in PDAC. Show less
Schaick, G. van; Dominguez Vega, E.; Gstottner, C.; Berg-Verleg, J.H. van den; Schouten, O.; Akeroyd, M.; ... ; Franc, V. 2022
Cells are covered with a dense layer of carbohydrates, some of which are solely present on neoplastic cells. The so-called tumor-associated carbohydrate antigens (TACAs) are increasingly recognized... Show moreCells are covered with a dense layer of carbohydrates, some of which are solely present on neoplastic cells. The so-called tumor-associated carbohydrate antigens (TACAs) are increasingly recognized as promising targets for immunotherapy. These carbohydrates differ from those of the surrounding non-cancerous tissues and contribute to the malignant phenotype of the cancer cells by promoting proliferation, metastasis, and immunosuppression. However, due to tumor tissue heterogeneity and technological limitations, TACAs are insufficiently explored.Methods: A workflow was established to decode the colorectal cancer (CRC)-associated O-linked glycans from approximately 20,000 cell extracts. Extracts were obtained through laser capture microdissection of formalin fixed paraffin embedded tissues of both primary tumors and metastatic sites, and compared to healthy colon mucosa from the same patients. The released O-glycans were analyzed by porous graphitized carbon liquid chromatography-tandem mass spectrometry in negative ion mode.Results: Distinctive O-glycosylation features were found in cancerous, stromal and normal colon mucosal regions. Over 100 O-linked glycans were detected in cancerous regions with absence in normal mucosa. From those, six core 2 O-glycans were exclusively found in more than 33% of the cancers, carrying the terminal (sialyl-)Lewis(X/A) antigen. Moreover, two O-glycans were present in 72% of the analyzed cancers and 94% of the investigated cancers expressed at least one of these two O-glycans. In contrast, normal colon mucosa predominantly expressed core 3 O-glycans, carrying alpha 2-6-linked sialylation, (sulfo-)Lewis(X/A) and Sda antigens.Conclusion: In this study, we present a novel panel of highly specific TACAs, based upon differences in the glycomic profiles between CRC and healthy colon mucosa. These TACAs are promising new targets for development of innovative cancer immune target therapies and lay the foundation for the targeted treatment of CRC. Show less
As a lymphoid organ, the spleen hosts a wide range of immune cell populations, which not only remove blood-borne antigens, but also generate and regulate antigen-specific immune responses. In... Show moreAs a lymphoid organ, the spleen hosts a wide range of immune cell populations, which not only remove blood-borne antigens, but also generate and regulate antigen-specific immune responses. In particular, the splenic microenvironment has been demonstrated to play a prominent role in adaptive immune responses to enveloped viral infections and alloantigens. During both types of immunizations, antigen-specific immunoglobulins G (IgGs) have been characterized by the reduced amount of fucose present on N-linked glycans of the fragment crystallizable (Fc) region. These glycans are essential for mediating the induction of immune effector functions. Therefore, we hypothesized that a spleen may modulate humoral responses and serve as a preferential site for afucosylated IgG responses, which potentially play a role in immune thrombocytopenia (ITP) pathogenesis. To determine the role of the spleen in IgG-Fc glycosylation, we performed IgG subclass-specific liquid chromatography-mass spectrometry (LC-MS) analysis of Fc glycosylation in a large cohort of individuals splenectomized due to trauma, due to ITP, or spherocytosis. IgG-Fc fucosylation was consistently increased after splenectomy, while no effects for IgG-Fc galactosylation and sialylation were observed. An increase in IgG1- and IgG2/3-Fc fucosylation level upon splenectomy has been reported here for the first time, suggesting that immune responses occurring in the spleen may be particularly prone to generate afucosylated IgG responses. Surprisingly, the level of total IgG-Fc fucosylation was decreased in ITP patients compared to healthy controls. Overall, our results suggest a yet unrecognized role of the spleen in either the induction or maintenance of afucosylated IgG responses by B cells. Show less
Selke, P.; Bork, K.; Zhang, T.; Wuhrer, M.; Strauss, C.; Horstkorte, R.; Scheer, M. 2021
Meningiomas are the most common non-malignant intracranial tumors and prefer, like most tumors, anaerobic glycolysis for energy production (Warburg effect). This anaerobic glycolysis leads to an... Show moreMeningiomas are the most common non-malignant intracranial tumors and prefer, like most tumors, anaerobic glycolysis for energy production (Warburg effect). This anaerobic glycolysis leads to an increased synthesis of the metabolite methylglyoxal (MGO) or glyoxal (GO), which is known to react with amino groups of proteins. This reaction is called glycation, thereby building advanced glycation end products (AGEs). In this study, we investigated the influence of glycation on sialylation in two meningioma cell lines, representing the WHO grade I (BEN-MEN-1) and the WHO grade III (IOMM-Lee). In the benign meningioma cell line, glycation led to differences in expression of sialyltransferases (ST3GAL1/2/3/5/6, ST6GAL1/2, ST6GALNAC2/6, and ST8SIA1/2), which are known to play a role in tumor progression. We could show that glycation of BEN-MEN-1 cells led to decreased expression of ST3Gal5. This resulted in decreased synthesis of the ganglioside GM3, the product of ST3Gal5. In the malignant meningioma cell line, we observed changes in expression of sialyltransferases (ST3GAL1/2/3, ST6GALNAC5, and ST8SIA1) after glycation, which correlates with less aggressive behavior. Show less
