Endogenous glucuronyltransferase activity of LARGE or LARGE2 required for functional modification of α-dystroglycan in cells and tissues

Kei-ichiro Inamori; Tobias Willer; Yuji Hara; David Venzke; Mary E Anderson; Nigel F Clarke; Pascale Guicheney; Carsten G Bönnemann; Steven A Moore; Kevin P Campbell

doi:10.1074/jbc.M114.597831

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Endogenous glucuronyltransferase activity of LARGE or LARGE2 required for functional modification of α-dystroglycan in cells and tissues

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Endogenous glucuronyltransferase activity of LARGE or LARGE2 required for functional modification of α-dystroglycan in cells and tissues

Kei-ichiro Inamori, Tobias Willer, Yuji Hara, David Venzke, Mary E Anderson, Nigel F Clarke, Pascale Guicheney, Carsten G Bönnemann, Steven A Moore and Kevin P Campbell

The Journal of biological chemistry, Vol.289(41), pp.28138-28148

10/10/2014

DOI: 10.1074/jbc.M114.597831

PMCID: PMC4192470

PMID: 25138275

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Abstract

Mutations in the LARGE gene have been identified in congenital muscular dystrophy (CMD) patients with brain abnormalities. Both LARGE and its paralog, LARGE2 (also referred to as GYLTL1B) are bifunctional glycosyltransferases with xylosyltransferase (Xyl-T) and glucuronyltransferase (GlcA-T) activities, and are capable of forming polymers consisting of [-3Xyl-α1,3GlcAβ1-] repeats. LARGE-dependent modification of α-dystroglycan (α-DG) with these polysaccharides is essential for the ability of α-DG to act as a receptor for ligands in the extracellular matrix. Here we report on the endogenous enzymatic activities of LARGE and LARGE2 in mice and humans, using a newly developed assay for GlcA-T activity. We show that normal mouse and human cultured cells have endogenous LARGE GlcA-T, and that this activity is absent in cells from the Large(myd) (Large-deficient) mouse model of muscular dystrophy, as well as in cells from CMD patients with mutations in the LARGE gene. We also demonstrate that GlcA-T activity is significant in the brain, heart, and skeletal muscle of wild-type and Large2(-/-) mice, but negligible in the corresponding tissues of the Large(myd) mice. Notably, GlcA-T activity is substantial, though reduced, in the kidneys of both the Large(myd) and Large2(-/-) mice, consistent with the observation of α-DG/laminin binding in these contexts. This study is the first to test LARGE activity in samples as small as cryosections and, moreover, provides the first direct evidence that not only LARGE, but also LARGE2, is vital to effective functional modification of α-DG in vivo.

Fibroblasts - enzymology

Kidney - pathology

Humans

Kidney - enzymology

Brain - enzymology

Dystroglycans - metabolism

N-Acetylglucosaminyltransferases - genetics

Muscular Dystrophies - genetics

Muscular Dystrophies - enzymology

Female

Glycosyltransferases - genetics

Binding Sites

Child

Disease Models, Animal

Muscle, Skeletal - enzymology

Cells, Cultured

Gene Expression Regulation

Myocardium - pathology

Enzyme Assays

Fibroblasts - pathology

Muscular Dystrophies - pathology

Organ Specificity

N-Acetylglucosaminyltransferases - metabolism

Mice, Knockout

Laminin - genetics

Glycosyltransferases - metabolism

Myocardium - enzymology

Animals

Brain - pathology

Protein Binding

Dystroglycans - genetics

Mice

Laminin - metabolism

Muscle, Skeletal - pathology

Details

Title: Subtitle: Endogenous glucuronyltransferase activity of LARGE or LARGE2 required for functional modification of α-dystroglycan in cells and tissues
Creators: Kei-ichiro Inamori - From the Howard Hughes Medical Institute, Department of Molecular Physiology and Biophysics, Department of Neurology, Department of Internal Medicine, University of Iowa Roy J. and Lucille A. Carver College of Medicine, Iowa City, Iowa 52242-1101, Division of Glycopathology, Institute of Molecular Biomembrane and Glycobiology, Tohoku Pharmaceutical University, Sendai, Miyagi 981-8558, Japan
Tobias Willer - From the Howard Hughes Medical Institute, Department of Molecular Physiology and Biophysics, Department of Neurology, Department of Internal Medicine, University of Iowa Roy J. and Lucille A. Carver College of Medicine, Iowa City, Iowa 52242-1101
Yuji Hara - From the Howard Hughes Medical Institute, Department of Molecular Physiology and Biophysics, Department of Neurology, Department of Internal Medicine, University of Iowa Roy J. and Lucille A. Carver College of Medicine, Iowa City, Iowa 52242-1101
David Venzke - From the Howard Hughes Medical Institute, Department of Molecular Physiology and Biophysics, Department of Neurology, Department of Internal Medicine, University of Iowa Roy J. and Lucille A. Carver College of Medicine, Iowa City, Iowa 52242-1101
Mary E Anderson - From the Howard Hughes Medical Institute, Department of Molecular Physiology and Biophysics, Department of Neurology, Department of Internal Medicine, University of Iowa Roy J. and Lucille A. Carver College of Medicine, Iowa City, Iowa 52242-1101
Nigel F Clarke - Institute for Neuroscience and Muscle Research, The Children's Hospital at Westmead, University of Sydney, Sydney, Australia
Pascale Guicheney - Inserm, U1166, Faculté de Médecine Pierre et Marie Curie, Institute of Cardiometabolism and Nutrition, ICAN, Paris, France, Sorbonne Universités, UPMC Univ Paris 06, UMR_S1166, Paris, France
Carsten G Bönnemann - Neurogenetics Branch, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892
Steven A Moore - Department of Pathology, University of Iowa Roy J. and Lucille A. Carver College of Medicine, Iowa City, Iowa 52242
Kevin P Campbell - From the Howard Hughes Medical Institute, Department of Molecular Physiology and Biophysics, Department of Neurology, Department of Internal Medicine, University of Iowa Roy J. and Lucille A. Carver College of Medicine, Iowa City, Iowa 52242-1101, kevin-campbell@uiowa.edu
Resource Type: Journal article
Publication Details: The Journal of biological chemistry, Vol.289(41), pp.28138-28148
DOI: 10.1074/jbc.M114.597831
PMID: 25138275
PMCID: PMC4192470
NLM abbreviation: J Biol Chem
ISSN: 0021-9258
eISSN: 1083-351X
Publisher: United States
Grant note: U54 NS053672 / NINDS NIH HHS 1 U54 NS053672 / NINDS NIH HHS 1 RC2 NS069521-01 / NINDS NIH HHS Howard Hughes Medical Institute RC2 NS069521 / NINDS NIH HHS
Language: English
Date published: 10/10/2014
Academic Unit: Neurology; Molecular Physiology and Biophysics; Pathology; Iowa Neuroscience Institute
Record Identifier: 9984020502402771

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