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Cells 2018, 7(5), 44; https://doi.org/10.3390/cells7050044

OGT (O-GlcNAc Transferase) Selectively Modifies Multiple Residues Unique to Lamin A

1
Department of Cell Biology, Johns Hopkins University School of Medicine, 725 North Wolfe Street, Baltimore, MD 21205, USA
2
Department of Chemistry, University of Virginia, Charlottesville, VA 22904, USA
3
Department of Nutrition, Institute of Basic Medical Sciences, University of Oslo, 0317 Oslo, Norway
4
Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
5
Department of Genetics, Institute of Life Sciences, Hebrew University of Jerusalem, Givat Ram Jerusalem 91904, Israel
6
Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway
7
Department of Pathology, University of Virginia, Charlottesville, VA 22904, USA
These authors contributed equally to this work.
Current address: The Laboratory of Cellular and Structural Biology, The Rockefeller University, New York, NY 10065, USA.
§
Current address: Department of Biochemistry, Duke University School of Medicine, Durham, NC 27708, USA.
Current address: Global Oral Care R&D, Johnson & Johnson, Skillman, NJ 08558, USA.
*
Author to whom correspondence should be addressed.
Received: 16 April 2018 / Revised: 11 May 2018 / Accepted: 14 May 2018 / Published: 17 May 2018
(This article belongs to the Collection Lamins and Laminopathies)
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Abstract

The LMNA gene encodes lamins A and C with key roles in nuclear structure, signaling, gene regulation, and genome integrity. Mutations in LMNA cause over 12 diseases (‘laminopathies’). Lamins A and C are identical for their first 566 residues. However, they form separate filaments in vivo, with apparently distinct roles. We report that lamin A is β-O-linked N-acetylglucosamine-(O-GlcNAc)-modified in human hepatoma (Huh7) cells and in mouse liver. In vitro assays with purified O-GlcNAc transferase (OGT) enzyme showed robust O-GlcNAcylation of recombinant mature lamin A tails (residues 385–646), with no detectable modification of lamin B1, lamin C, or ‘progerin’ (Δ50) tails. Using mass spectrometry, we identified 11 O-GlcNAc sites in a ‘sweet spot’ unique to lamin A, with up to seven sugars per peptide. Most sites were unpredicted by current algorithms. Double-mutant (S612A/T643A) lamin A tails were still robustly O-GlcNAc-modified at seven sites. By contrast, O-GlcNAcylation was undetectable on tails bearing deletion Δ50, which causes Hutchinson–Gilford progeria syndrome, and greatly reduced by deletion Δ35. We conclude that residues deleted in progeria are required for substrate recognition and/or modification by OGT in vitro. Interestingly, deletion Δ35, which does not remove the majority of identified O-GlcNAc sites, does remove potential OGT-association motifs (lamin A residues 622–625 and 639–645) homologous to that in mouse Tet1. These biochemical results are significant because they identify a novel molecular pathway that may profoundly influence lamin A function. The hypothesis that lamin A is selectively regulated by OGT warrants future testing in vivo, along with two predictions: genetic variants may contribute to disease by perturbing OGT-dependent regulation, and nutrient or other stresses might cause OGT to misregulate wildtype lamin A. View Full-Text
Keywords: lamin; nuclear lamina; O-GlcNAcylation; O-linked N-acetylglucosamine (O-GlcNAc) Transferase (OGT) lamin; nuclear lamina; O-GlcNAcylation; O-linked N-acetylglucosamine (O-GlcNAc) Transferase (OGT)
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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Simon, D.N.; Wriston, A.; Fan, Q.; Shabanowitz, J.; Florwick, A.; Dharmaraj, T.; Peterson, S.B.; Gruenbaum, Y.; Carlson, C.R.; Grønning-Wang, L.M.; Hunt, D.F.; Wilson, K.L. OGT (O-GlcNAc Transferase) Selectively Modifies Multiple Residues Unique to Lamin A. Cells 2018, 7, 44.

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