1. Biochemistry and Chemical Biology
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The glucuronyltransferase B4GAT1 is required for initiation of LARGE-mediated α-dystroglycan functional glycosylation

  1. Tobias Willer
  2. Kei-ichiro Inamori
  3. David Venzke
  4. Corinne Harvey
  5. Greg Morgensen
  6. Yuji Hara
  7. Daniel Beltrán Valero de Bernabé
  8. Liping Yu
  9. Kevin M Wright
  10. Kevin P Campbell  Is a corresponding author
  1. Howard Hughes Medical Institute, University of Iowa, Carver College of Medicine, United States
  2. Tohoku Pharmaceutical University, Japan
  3. Graduate School of Engineering, Kyoto University, Japan
  4. University of Iowa, Carver College of Medicine, United States
  5. Oregon Health and Science University, United States
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Cite this article as: eLife 2014;3:e03941 doi: 10.7554/eLife.03941

Abstract

Dystroglycan is a cell membrane receptor that organizes the basement membrane by binding ligands in the extracellular matrix. Proper glycosylation of the α-dystroglycan (α-DG) subunit is essential for these activities, and lack thereof results in neuromuscular disease. Currently, neither the glycan synthesis pathway nor the roles of many known or putative glycosyltransferases that are essential for this process are well understood. Here we show that FKRP, FKTN, TMEM5 and B4GAT1 (formerly known as B3GNT1) localize to the Golgi and contribute to the O-mannosyl post-phosphorylation modification of α-DG. Moreover, we assigned B4GAT1 a function as a xylose β1,4-glucuronyltransferase. Nuclear magnetic resonance studies confirmed that a glucuronic acid β1,4-xylose disaccharide synthesized by B4GAT1 acts as an acceptor primer that can be elongated by LARGE with the ligand-binding heteropolysaccharide. Our findings greatly broaden the understanding of α-DG glycosylation and provide mechanistic insight into why mutations in B4GAT1 disrupt dystroglycan function and cause disease.

Article and author information

Author details

  1. Tobias Willer

    Howard Hughes Medical Institute, University of Iowa, Carver College of Medicine, Iowa City, United States
    Competing interests
    The authors declare that no competing interests exist.

  2. Kei-ichiro Inamori

    Tohoku Pharmaceutical University, Komatsushima, Japan
    Competing interests
    The authors declare that no competing interests exist.

  3. David Venzke

    Howard Hughes Medical Institute, University of Iowa, Carver College of Medicine, Iowa City, United States
    Competing interests
    The authors declare that no competing interests exist.

  4. Corinne Harvey

    Howard Hughes Medical Institute, University of Iowa, Carver College of Medicine, Iowa City, United States
    Competing interests
    The authors declare that no competing interests exist.

  5. Greg Morgensen

    Howard Hughes Medical Institute, University of Iowa, Carver College of Medicine, Iowa City, United States
    Competing interests
    The authors declare that no competing interests exist.

  6. Yuji Hara

    Graduate School of Engineering, Kyoto University, Kyoto, Japan
    Competing interests
    The authors declare that no competing interests exist.

  7. Daniel Beltrán Valero de Bernabé

    Howard Hughes Medical Institute, University of Iowa, Carver College of Medicine, Iowa City, United States
    Competing interests
    The authors declare that no competing interests exist.

  8. Liping Yu

    University of Iowa, Carver College of Medicine, Iowa City, United States
    Competing interests
    The authors declare that no competing interests exist.

  9. Kevin M Wright

    Oregon Health and Science University, Portland, United States
    Competing interests
    The authors declare that no competing interests exist.

  10. Kevin P Campbell

    Howard Hughes Medical Institute, University of Iowa, Carver College of Medicine, Iowa City, United States
    For correspondence
    kevin-campbell@uiowa.edu
    Competing interests
    The authors declare that no competing interests exist.

Ethics

Animal experimentation: Animal care, ethical usage and procedures were approved and performed in accordance with the standards set forth by the National Institutes of Health and the Animal Care Use and Review Committee at the University of Iowa (protocol #4081122). At the University of Iowa all mice are socially housed (unless single housing is required) under specific pathogen-free conditions in an AAALAC accredited animal facility. Housing conditions are as specified in the Guide for the Care and Use of Laboratory Animals (NRC). Mice are housed on Thoren brand, HEPA filtered ventilated racks, in solid bottom cages with mixed paper bedding. A standard 12/12-h light/dark cycle was used. Standard rodent chow (or special diet if required) and water is available ad libitum.

Reviewing Editor

  1. Suzanne R Pfeffer, Stanford University, United States

Publication history

  1. Received: July 9, 2014
  2. Accepted: October 1, 2014
  3. Accepted Manuscript published: October 3, 2014 (version 1)
  4. Version of Record published: November 11, 2014 (version 2)

Copyright

© 2014, Willer et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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    B4GAT1 is the priming enzyme for the LARGE-dependent functional glycosylation of α-dystroglycan

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    Research Article Updated

    Recent studies demonstrated that mutations in B3GNT1, an enzyme proposed to be involved in poly-N-acetyllactosamine synthesis, were causal for congenital muscular dystrophy with hypoglycosylation of α-dystroglycan (secondary dystroglycanopathies). Since defects in the O-mannosylation protein glycosylation pathway are primarily responsible for dystroglycanopathies and with no established O-mannose initiated structures containing a β3 linked GlcNAc known, we biochemically interrogated this human enzyme. Here we report this enzyme is not a β-1,3-N-acetylglucosaminyltransferase with catalytic activity towards β-galactose but rather a β-1,4-glucuronyltransferase, designated B4GAT1, towards both α- and β-anomers of xylose. The dual-activity LARGE enzyme is capable of extending products of B4GAT1 and we provide experimental evidence that B4GAT1 is the priming enzyme for LARGE. Our results further define the functional O-mannosylated glycan structure and indicate that B4GAT1 is involved in the initiation of the LARGE-dependent repeating disaccharide that is necessary for extracellular matrix protein binding to O-mannosylated α-dystroglycan that is lacking in secondary dystroglycanopathies.

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