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Dystroglycan glycosylation and its role in alpha-dystroglycanopathies.

ABSTRACT: Glycosylation is the most common post-translational modification of proteins. The protein sequence data suggested that more than half of all proteins produced in mammalian cells are glycoproteins. Glycans of secreted glycoproteins affect many protein properties such as solubility, stability, protease sensitivity, and polarity, while glycans on cell surface glycoproteins are involved in various cellular functions including cell-cell and cell-matrix interactions during embryogenesis, immune reactions, and tumor development. The past decade of research on glycan function has revealed the etiology of a growing number of human genetic diseases with aberrant glycan formation. This review focuses upon the involvement of O-mannosylglycan in the molecular and cellular mechanisms of muscular dystrophies. Advances in glycobiology are expected to result in a better understanding and in improved treatments of a class of muscular dystrophies called alpha-dystroglycanopathies.

SUBMITTER: Endo T

PROVIDER: S-EPMC2949307 | biostudies-literature | 2007 Dec

REPOSITORIES: biostudies-literature

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Publications

Dystroglycan glycosylation and its role in alpha-dystroglycanopathies.

Endo T T

Acta myologica : myopathies and cardiomyopathies : official journal of the Mediterranean Society of Myology 20071201 3

Glycosylation is the most common post-translational modification of proteins. The protein sequence data suggested that more than half of all proteins produced in mammalian cells are glycoproteins. Glycans of secreted glycoproteins affect many protein properties such as solubility, stability, protease sensitivity, and polarity, while glycans on cell surface glycoproteins are involved in various cellular functions including cell-cell and cell-matrix interactions during embryogenesis, immune reacti ...[more]

PMID: 18646566

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Project description:?-dystroglycan (?-DG) is a peripheral membrane protein that is an integral component of the dystrophin-glycoprotein complex. In an inherited subset of muscular dystrophies known as dystroglycanopathies, ?-DG has reduced glycosylation which results in lower affinity binding to several extracellular matrix proteins including laminins. The glycosylation status of ?-DG is normally assessed by the binding of the ?-DG antibody IIH6 to a specific glycan epitope on ?-DG involved in laminin binding. Immunocytochemistry and immunoblotting are two of the most widely used methods to detect the amount of ?-DG glycosylation in muscle. While the interpretation of the presence or absence of the epitope on muscle using these techniques is straightforward, the assessment of a mild defect can be challenging. In this study, flow cytometry was used to compare the amount of IIH6-reactive glycans in fibroblasts from dystroglycanopathy patients with defects in genes known to cause ?-DG hypoglycosylation to the amount in fibroblasts from healthy and pathological control subjects. A total of twenty one dystroglycanopathy patient fibroblasts were assessed, as well as fibroblasts from three healthy controls and seven pathological controls. Control fibroblasts have clearly detectable amounts of IIH6-reactive glycans, and there is a significant difference in the amount of this glycosylation, as measured by the mean fluorescence intensity of an antibody recognising the epitope and the percentage of cells positive for the epitope, between these controls and dystroglycanopathy patient fibroblasts (p<0.0001 for both). Our results indicate that the amount of ?-DG glycosylation in patient fibroblasts is comparable to that in patient skeletal muscle. This method could complement existing immunohistochemical assays in skeletal muscle as it is quantitative and simple to perform, and could be used when a muscle biopsy is not available. This test could also be used to assess the pathogenicity of variants of unknown significance in genes involved in dystroglycanopathies.

Project description:The Congenital Disorders of Glycosylation (CDG) are an expanding group of genetic disorders which encompass a spectrum of glycosylation defects of protein and lipids, including N- & O-linked defects and among the latter are the muscular dystroglycanopathies (MD). Initial screening of CDG is usually based on the investigation of the glycoproteins transferrin, and/or apolipoprotein CIII. These biomarkers do not always detect complex or subtle defects present in older patients, therefore there is a need to investigate additional glycoproteins in some cases. We describe a sensitive 2D-Differential Gel Electrophoresis (DIGE) method that provides a global analysis of the serum glycoproteome. Patient samples from PMM2-CDG (n = 5), CDG-II (n = 7), MD and known complex N- & O-linked glycosylation defects (n = 3) were analysed by 2D DIGE. Using this technique we demonstrated characteristic changes in mass and charge in PMM2-CDG and in charge in CDG-II for ?1-antitrypsin, ?1-antichymotrypsin, ?2-HS-glycoprotein, ceruloplasmin, and ?1-acid glycoproteins 1&2. Analysis of the samples with known N- & O-linked defects identified a lower molecular weight glycoform of C1-esterase inhibitor that was not observed in the N-linked glycosylation disorders indicating the change is likely due to affected O-glycosylation. In addition, we could identify abnormal serum glycoproteins in LARGE and B3GALNT2-deficient muscular dystrophies. The results demonstrate that the glycoform pattern is varied for some CDG patients not all glycoproteins are consistently affected and analysis of more than one protein in complex cases is warranted. 2D DIGE is an ideal method to investigate the global glycoproteome and is a potentially powerful tool and secondary test for aiding the complex diagnosis and sub classification of CDG. The technique has further potential in monitoring patients for future treatment strategies. In an era of shifting emphasis from gel- to mass-spectral based proteomics techniques, we demonstrate that 2D-DIGE remains a powerful method for studying global changes in post-translational modifications of proteins.

Dataset Information

Dystroglycan glycosylation and its role in alpha-dystroglycanopathies.

Publications

Dystroglycan glycosylation and its role in alpha-dystroglycanopathies.

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