Protein O-Mannosylation in the Murine Brain: Occurrence of Mono-O-Mannosyl Glycans and Identification of New Substrates.
Protein O-mannosylation is a post-translational modification essential for correct development of mammals. In humans, deficient O-mannosylation results in severe congenital muscular dystrophies often associated with impaired brain and eye development. Although various O-mannosylated proteins have been identified in the recent years, the distribution of O-mannosyl glycans in the mammalian brain and target proteins are still not well defined. In the present study, rabbit monoclonal antibodies directed against the O-mannosylated peptide YAT(α1-Man)AV were generated. Detailed characterization of clone RKU-1-3-5 revealed that this monoclonal antibody recognizes O-linked mannose also in different peptide and protein contexts. Using this tool, we observed that mono-O-mannosyl glycans occur ubiquitously throughout the murine brain but are especially enriched at inhibitory GABAergic neurons and at the perineural nets. Using a mass spectrometry-based approach, we further identified glycoproteins from the murine brain that bear single O-mannose residues. Among the candidates identified are members of the cadherin and plexin superfamilies and the perineural net protein neurocan. In addition, we identified neurexin 3, a cell adhesion protein involved in synaptic plasticity, and inter-alpha-trypsin inhibitor 5, a protease inhibitor important in stabilizing the extracellular matrix, as new O-mannosylated glycoproteins.
Centre for Organismal Studies, Department of Cell Chemistry, Heidelberg University, Heidelberg, Germany.
Markus F
MF
Bartels
Centre for Organismal Studies, Department of Cell Chemistry, Heidelberg University, Heidelberg, Germany.|Center for Molecular Biology Heidelberg (ZMBH), Core Facility for Mass Spectrometry and Proteomics, Heidelberg University, Heidelberg, Germany.
Patrick R
PR
Winterhalter
Gesellschaft zur Förderung der Analytischen Wissenschaften e.V., ISAS-Leibniz Institute for Analytical Sciences, Dortmund, Germany.
Jin
J
Yu
Glycosciences Laboratory, Department of Medicine, Imperial College London, Hammersmith Campus, London, United Kingdom.
Yan
Y
Liu
Centre for Organismal Studies, Department of Cell Chemistry, Heidelberg University, Heidelberg, Germany.
Mark
M
Lommel
Centre for Organismal Studies, Department of Animal Molecular Physiology, Heidelberg University, Heidelberg, Germany.
Frank
F
Möhrlen
Institute for Human Performance, Department of Neuroscience and Physiology, Upstate Medical University, New York, United States of America.
Huaiyu
H
Hu
Glycosciences Laboratory, Department of Medicine, Imperial College London, Hammersmith Campus, London, United Kingdom.
Ten
T
Feizi
Gesellschaft zur Förderung der Analytischen Wissenschaften e.V., ISAS-Leibniz Institute for Analytical Sciences, Dortmund, Germany.
Ulrika
U
Westerlind
Center for Molecular Biology Heidelberg (ZMBH), Core Facility for Mass Spectrometry and Proteomics, Heidelberg University, Heidelberg, Germany.
Thomas
T
Ruppert
Centre for Organismal Studies, Department of Cell Chemistry, Heidelberg University, Heidelberg, Germany.
Sabine
S
Strahl
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D059330:GABAergic Neurons / Q000378:metabolism; D006023:Glycoproteins / Q000737:chemistry* / Q000378:metabolism*; D008358:Mannose / Q000378:metabolism*; D051379:Mice; D011134:Polysaccharides / Q000737:chemistry* / Q000378:metabolism*; D011499:Protein Processing, Post-Translational
D006023:Glycoproteins; D011134:Polysaccharides; D008358:Mannose