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The fungal cell wall occupies a central place in the interaction between fungi and their environment. This study focuses on the role of the putative polysaccharide synthase Cps1 in the physiology, development and virulence of the grey mold-causing agent Botrytis cinerea. Deletion of the Bccps1 gene does not affect the germination of the conidia (asexual spores) or the early mycelial development, but it perturbs hyphal expansion after 24 h, revealing a two-phase hyphal development that has not been reported so far. It causes a severe reduction of mycelial growth in a solid medium and modifies hyphal aggregation into pellets in liquid cultures. It strongly impairs plant penetration, plant colonization and the formation of sclerotia (survival structures). Loss of the BcCps1 protein associates with a decrease in glucans and glycoproteins in the fungus cell wall and the up-accumulation of 132 proteins in the mutant's exoproteome, among which are fungal cell wall enzymes. This is accompanied by an increased fragility of the mutant mycelium, an increased sensitivity to some environmental stresses and a reduced adhesion to plant surface. Taken together, the results support a significant role of Cps1 in the cell wall biology of B. cinerea. [ABSTRACT FROM AUTHOR]

The fungal cell wall occupies a central place in the interaction between fungi and their environment. This study focuses on the role of the putative polysaccharide synthase Cps1 in the physiology, development and virulence of the grey mold-causing agent Botrytis cinerea. Deletion of the Bccps1 gene does not affect the germination of the conidia (asexual spores) or the early mycelial development, but it perturbs hyphal expansion after 24 h, revealing a two-phase hyphal development that has not been reported so far. It causes a severe reduction of mycelial growth in a solid medium and modifies hyphal aggregation into pellets in liquid cultures. It strongly impairs plant penetration, plant colonization and the formation of sclerotia (survival structures). Loss of the BcCps1 protein associates with a decrease in glucans and glycoproteins in the fungus cell wall and the up-accumulation of 132 proteins in the mutant’s exoproteome, among which are fungal cell wall enzymes. This is accompanied by an increased fragility of the mutant mycelium, an increased sensitivity to some environmental stresses and a reduced adhesion to plant surface. Taken together, the results support a significant role of Cps1 in the cell wall biology of B. cinerea.

The high-prevalence blood group antigen, Sd a , had been puzzling blood bankers and transfusionists for at least a decade when it was reported in 1967. The characteristic mix of agglutinates and free red blood cells (RBCs), caused by anti-Sd a , is seen with the RBCs from 90 percent of individuals of European descent. However, only 2-4 percent of individuals are truly Sd(a-) and may produce anti-Sd a . The antibodies, generally considered insignificant, may cause hemolytic transfusion reactions with high-expressing Sd(a+) RBCs (e.g., the unusual Cad phenotype, which can also be polyagglutinable). The Sd a glycan, GalNAcβ1-4(NeuAcα2-3)Gal-R, is produced in the gastrointestinal and urinary systems, while its origin on RBCs is more controversial. According to current theory, Sd a is likely to be passively adsorbed in low amounts, except in Cad individuals, where it has been found on erythroid proteins and at higher levels. The long-standing hypothesis that B4GALNT2 encodes the Sd a synthase was confirmed in 2019, since homozygosity for a variant allele with rs7224888:C produces a non-functional enzyme associated with most cases of the Sd(a-) phenotype. Thereby, the SID blood group system was acknowledged as number 038 by the International Society of Blood Transfusion. Although the genetic background of Sd(a-) was settled, questions remain. The genetic background of the Cad phenotype has not yet been determined, and the source of the RBC-carried Sd a is unknown. Furthermore, the interest of Sd a stretches beyond transfusion medicine. Some tantalizing examples are lowered antigen levels in malignant tissue compared with normal tissue and interference with infectious agents like Escherichia coli , influenza virus, and malaria parasites., (© 2023 L. Stenfelt et al., published by Sciendo.)

Most proteins are modified by glycans, which can modulate the biological properties and functions of glycoproteins. The major glycans can be classified into N-glycans and O-glycans according to their glycan-peptide linkage. This review will provide an overview of the O-mannosyl glycans, one subtype of O-glycans. Originally, O-mannosyl glycan was only known to be present on a limited number of glycoproteins, especially α-dystroglycan (α-DG). However, once a clear relationship was established between O-mannosyl glycan and the pathological mechanisms of some congenital muscular dystrophies in humans, research on the biochemistry and pathology of O-mannosyl glycans has been expanding. Because α-DG glycosylation is defective in congenital muscular dystrophies, which also feature abnormal neuronal migration, these disorders are collectively called α-dystroglycanopathies. In this article, I will describe the structure, biosynthesis and pathology of O-mannosyl glycans. [ABSTRACT FROM AUTHOR]

Aberrant glycosylation may promote tumor invasion and metastasis. To investigate whether microRNA (miRNA) is involved in glycosylation-related metastasis, we examined the role of let-7c, a well-known tumor-suppressor miRNA, in glycosylation in murine hepatocarcinoma cell lines Hca-F and Hca-P. We found that let-7c level was higher in Hca-P cells (with lower lymphatic metastasis potential) than in Hca-F cells (with higher lymphatic metastasis potential). Overexpression of let-7c decreased hyper-N-glycosylation of Hca-F cells and repressed their metastatic and invasive ability. Mannoside acetylglucosaminyltransferase 4, isoenzyme A (Mgat4a) is a key glycosyltransferase in the pathway of synthesizing complex N-glycans. Bioinformatics analysis indicates that Mgat4a may be a target of let-7c, which has been verified by dual-luciferase reporter gene assay. Furthermore, the anti-metastatic effect of overexpressed let-7c is similar to that of Mgat4a siRNAs transfection. Hence, our results suggest that let-7c may inhibit the metastatic ability of Hca-F cells, at least partially, via repressing Mgat4a activity. [ABSTRACT FROM AUTHOR]

Abstract: The surface of Trypanosoma cruzi is covered by a dense glycocalix which is characteristic of each stage of the life cycle. Its composition and complexity depend mainly on mucin-like proteins. A remarkable feature of O-glycan biosynthesis in trypanosomes is that it initiates with the addition of a GlcNAc instead of the GalNAc residue that is commonly used in vertebrate mucins. The fact that the interplay between trans-sialidase and mucin is crucial for pathogenesis, and both families have stage-specific members is also remarkable. Recently the enzyme that transfers the first GlcNAc from UDP-GlcNAc to a serine or threonine residue was kinetically characterized. The relevance of this enzyme is evidenced by its role as catalyzer of the first step in O-glycosylation. In this paper we describe how this gene is expressed differentially along the life cycle with a pattern that is very similar to that of trans-sialidases. Its localization was determined, showing that the protein predicted to be in the Golgi apparatus is also present in reservosomes. Finally our results indicate that this enzyme, when overexpressed, enhances T. cruzi infectivity. [Copyright &y& Elsevier]

The article focuses on developments in the enzymatic synthesis of oligosaccharides and neoglycoconjugates. Sufficient quantities of structurally defined oligosaccharides are of limited availability by traditional purification method to elucidate the biological functions of oligosaccharides. Thus, chemical and enzymatic syntheses of oligosaccharides are becoming increasingly important in glycobiology and glycotechnology.

The CNS consists of neuronal and glial cells generated from common neural progenitor cells during development. Cellular events for neural progenitor cells, such as proliferation and differentiation, are regulated by multiple intrinsic and extrinsic cell signals. Although much is known on the importance of the proteinousfactors in regulating the fate of neural progenitor cells, the involvement of other molecules such as gangliosides, sialic acid-containing glycosphingolipids, remains to be clarified. To elucidate the biological functions of gangliosides in neural progenitor cells, we transfected an immortalized neural progenitor cell line, C17.2, which does not express GD3 ganglioside, with a fusion protein of GD3-synthase (ST-II) and enhanced green fluorescent protein (ST-II-EGFP). Analysis of the ST-II transfectants revealed the ectopic expression of b- and c-series gangliosides. In the ST-II transfectants, proliferation induced by epidermal growth factor (EGF) was severely retarded. EGF-induced proliferation of C17.2 cells was dependent on the Ras-mitogen-activated protein kinase (Ras-MAPK) pathway, and the EGF-induced activation of this pathway was significantly repressed in the transfectants. Thus, ST-II overexpression retarded proliferation of C17.2 cells via repression of the Ras-MAPK pathway. The result supports the concept that gangliosides may play an important role in regulating the proliferation of neural progenitor cells. [ABSTRACT FROM AUTHOR]

Purpose: To study the structural alterations of asparagine-linked sugar chains (N-glycans) on urine fibronectin (Fn) from bladder cancer (BCa) patients and its enzymatic mechanism. Methods: Eight pairs of urine samples from eight BCa patients pre-operation and 3 months post-operation (which proved to be normal) were collected, and the Fn in the urine samples was purified with an anti-Fn antibody affinity column. Different lectins labeled with horseradish peroxidase (HRP) were used as probes to bind the glycans of purified Fn immobilized on membrane. Enhanced chemiluminescence (ECL) reagent was adopted to estimate the activity of the bound HRP as a measure of the binding affinity of the Fn glycans to lectins, and expressed as luminescent light units (LLU). The enzymatic mechanism of the structural alteration of N-glycans in BCa Fn was studied by determination of GnT activities using the HPLC method and fluorescent-labeled substrate. Results: The mean LLU of BCa Fn was only 18.1% of the normal samples when Con A-HRP were used as probes, while the mean LLU of the BCa group was 3.34 times and 3.26 times higher than normal for the DSA-HRP and WGA-HRP probes, respectively. The individual data of the patients did not overlap between the BCa sample and normal counterparts, indicating that the positive rates were 100%, regardless of which lectin-HRP was used. These results reveal that the antennary number and bisecting GlcNAc structure are increased in the N-glycans of urine Fn from BCa assessed according to the binding specificity of ConA, DSA, and WGA. In addition, the binding affinities of urine Fn with DSA and WGA were correlated to pathological stage, and the affinity of Fn with WGA was also correlated with pathological grade. The results of GnT determination showed that GnT-III, IV, and V in BCa tissues increased by 34.0, 18.1, and 1.6 times, respectively, in normal bladder tissues which were at least 5 cm away from the BCa of the same bladder. These findings were compatible with the structural changes of N-glycans in BCa Fn, since GnT-III and GnT-IV/V are responsible for the synthesis of bisecting GlcNAc and the increase of antennary number in N-glycans, respectively. Conclusions: (1) The highest elevation of GnT-III and the close relationship between the WGA binding of BCa Fn with the pathological stage and grade of BCa indicate that the increase of bisecting GlcNAc in N-linked glycans contributes more to the malignant behavior of BCa than the increase of GnT-IV, GnT-V, and the antennary number. (2) The correlation of altered activities of bladder GnTs with the abnormal structures of urine Fn in BCa patients indicates that the urine Fn is synthesized in the bladder. (3) The lectin-HRP assay for analyzing the structure of N-glycans in urine Fn may be used as a simple and accurate diagnosis method for BCa in the future. [ABSTRACT FROM AUTHOR]

As an extension of a previous study (de Vries et al., 1995, J. Biol. Chem., 270, 8712–8722) the acceptor specificity of recombinant FucT VI, expressed in insect cells as soluble enzyme, and purified from the growth medium by affinity chromatography, was analyzed toward a broad panel of oligosaccharide and glycoprotein substrates. It was found that FucT V.1 effectively utilizes any type-2-chain based structure (Galβ1→4GlcNAc-R). Neutral as well as sialylated structures are fucosylated with high efficiency. To identify polar groups on acceptors that function in enzyme binding, deoxygenated substrate analogs were tested as acceptors. FucT VI had an absolute requirement for a hydroxyl at C-6 of galactose in addition to the accepting hydroxyl at C-3. Thus, FucT VI, although different from FucT III, IV, and V in acceptor properties, seems to bind the acceptor in a similar way. [ABSTRACT FROM PUBLISHER]

Chemoenzymatic oligosaccharide synthesis is attractive since it eliminates the tedious multistep protection-deprotection requirements of pure chemical synthesis. Chemoenzymatic synthesis using glycosyltransferases, however, requires not only the correct enzyme to control both regio- and stereospecificity, but also the glycosyl donor to provide the sugar that is added. This unit describes a simple synthesis of sugar-nucleoside diphosphates (sugar-NDPs), the type of glycosyl donor (e.g., UDP-Glc, UDP-Gal, ADP-Glc) required by most glycosyltransferases, by using a chemical coupling reaction in water. The preparation of sugar-NDPs by this method therefore does not require any skills in synthetic organic chemistry., (Copyright © 2013 John Wiley & Sons, Inc.)