Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Perspective
  • Published:

Tools, tactics and objectives to interrogate cellular roles of O-GlcNAc in disease

Abstract

The vast array of cell types of multicellular organisms must individually fine-tune their internal metabolism. One important metabolic and stress regulatory mechanism is the dynamic attachment/removal of glucose-derived sugar N-acetylglucosamine on proteins (O-GlcNAcylation). The number of proteins modified by O-GlcNAc is bewildering, with at least 7,000 sites in human cells. The outstanding challenge is determining how key O-GlcNAc sites regulate a target pathway amidst thousands of potential global sites. Innovative solutions are required to address this challenge in cell models and disease therapy. This Perspective shares critical suggestions for the O-GlcNAc field gleaned from the international O-GlcNAc community. Further, we summarize critical tools and tactics to enable newcomers to O-GlcNAc biology to drive innovation at the interface of metabolism and disease. The growing pace of O-GlcNAc research makes this a timely juncture to involve a wide array of scientists and new toolmakers to selectively approach the regulatory roles of O-GlcNAc in disease.

This is a preview of subscription content, access via your institution

Access options

Buy this article

Purchase on Springer Link

Instant access to full article PDF

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Cells use O-GlcNAc modifications to sense metabolites resulting from altered nutrient or stress levels, a process that alters protein behaviors and can lead to disease.
Fig. 2: Chemical biology tools reveal a plethora of O-GlcNAc cellular features.
Fig. 3: Site-specific chemical tools for O-GlcNAc studies.
Fig. 4: High levels of glucose and GlcNAc utilization in the brain make neurobiology a key area of study in the O-GlcNAc field.
Fig. 5: O-GlcNAc dynamics play extensive roles in epigenetic regulation, development and cancer.

Similar content being viewed by others

References

  1. Hart, G. W. Nutrient regulation of signaling and transcription. J. Biol. Chem. 294, 2211–2231 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Yang, X. & Qian, K. Protein O-GlcNAcylation: emerging mechanisms and functions. Nat. Rev. Mol. Cell Biol. 18, 452–465 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  3. Ferrer, C. M., Sodi, V. L. & Reginato, M. J. O-GlcNAcylation in cancer biology: linking metabolism and signaling. J. Mol. Biol. 428, 3282–3294 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Hanover, J. A., Chen, W. & Bond, M. R. O-GlcNAc in cancer: an oncometabolism-fueled vicious cycle. J. Bioenerg. Biomembr. 50, 155–173 (2018).

    Article  CAS  PubMed  Google Scholar 

  5. Ma, J. & Hart, G. W. Protein O-GlcNAcylation in diabetes and diabetic complications. Expert Rev. Proteom. 10, 365–380 (2013).

    Article  CAS  Google Scholar 

  6. Wang, A. C., Jensen, E. H., Rexach, J. E., Vinters, H. V. & Hsieh-Wilson, L. C. Loss of O-GlcNAc glycosylation in forebrain excitatory neurons induces neurodegeneration. Proc. Natl Acad. Sci. USA 113, 15120–15125 (2016). This paper used conditional knockout of OGT to precisely determine hippocampal neuron-specific roles for O-GlcNAc, especially neurodegeneration versus neuroprotection effects.

    Article  PubMed  PubMed Central  Google Scholar 

  7. Wheatley, E. G. et al. Neuronal O-GlcNAcylation improves cognitive function in the aged mouse brain. Curr. Biol. 29, 3359–3369 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Martinez, M. R., Dias, T. B., Natov, P. S. & Zachara, N. E. Stress-induced O-GlcNAcylation: an adaptive process of injured cells. Biochem. Soc. Trans. 45, 237–249 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Yang, Y. R. & Suh, P. G. O-GlcNAcylation in cellular functions and human diseases. Adv. Biol. Regul. 54, 68–73 (2014).

    Article  CAS  PubMed  Google Scholar 

  10. Slawson, C., Copeland, R. J. & Hart, G. W. O-GlcNAc signaling: a metabolic link between diabetes and cancer? Trends Biochem. Sci. 35, 547–555 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Wulff-Fuentes, E. et al. The human O-GlcNAcome database and meta-analysis. Sci. Data 8, 25 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Ma, J., Li, Y., Hou, C. & Wu, C. O-GlcNAcAtlas: a database of experimentally identified O-GlcNAc sites and proteins. Glycobiology 31, 719–723 (2021).

  13. Shafi, R. et al. The O-GlcNAc transferase gene resides on the X chromosome and is essential for embryonic stem cell viability and mouse ontogeny. Proc. Natl Acad. Sci. USA 97, 5735–5739 (2000).

    Article  PubMed Central  Google Scholar 

  14. O’Donnell, N., Zachara, N. E., Hart, G. W. & Marth, J. D. OGT-dependent X-chromosome-linked protein glycosylation is a requisite modification in somatic cell function and embryo viability. Mol. Cell. Biol. 24, 1680–1690 (2004).

    PubMed Central  Google Scholar 

  15. Yang, Y. R. et al. O-GlcNAcase is essential for embryonic development and maintenance of genomic stability. Aging Cell 11, 439–448 (2012).

    Article  CAS  PubMed  Google Scholar 

  16. Marshall, S., Nadeau, O. & Yamasaki, K. Dynamic actions of glucose and glucosamine on hexosamine biosynthesis in isolated adipocytes. J. Biol. Chem. 279, 35313–35319 (2004).

    Article  CAS  PubMed  Google Scholar 

  17. Zachara, N. E., Molina, H., Wong, K. Y., Pandey, A. & Hart, G. W. The dynamic stress-induced “O-GlcNAc-ome” highlights functions for O-GlcNAc in regulating DNA damage/repair and other cellular pathways. Amino Acids 40, 793–808 (2011).

    Article  CAS  PubMed  Google Scholar 

  18. Lu, L. et al. Distributive O-GlcNAcylation on the highly repetitive C-terminal domain of RNA polymerase II. Biochemistry 55, 1149–1158 (2016).

    Article  CAS  PubMed  Google Scholar 

  19. Tan, Z.-W. et al. O-GlcNAc regulates gene expression by controlling detained intron splicing. Nucleic Acids Res. 48, 5656–5669 (2020). This paper identifies an elegant mechanism for rapid re-balancing of OGT/OGA activity after O-GlcNAc disruption via O-GlcNAc-induced alternative splicing.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Miller, M. W., Caracciolo, M. R., Berlin, W. K. & Hanover, J. A. Phosphorylation and glycosylation of nucleoporins. Arch. Biochem. Biophys. 367, 51–60 (1999).

    Article  CAS  PubMed  Google Scholar 

  21. Eustice, M., Bond, M. R. & Hanover, J. A. O-GlcNAc cycling and the regulation of nucleocytoplasmic dynamics. Biochem. Soc. Trans. 45, 427–436 (2017).

    Article  CAS  PubMed  Google Scholar 

  22. Groenevelt, J. M., Corey, D. J. & Fehl, C. Chemical synthesis and biological applications of O-GlcNAcylated peptides and proteins. ChemBioChem 22, 1854–1870 (2021). This paper collects known synthetic methods for site-specific O-GlcNAc installation. Also see refs. 23 and 25 for thorough collections of O-GlcNAc detection tools and assays.

    Article  CAS  PubMed  Google Scholar 

  23. Gorelik, A. & van Aalten, D. M. F. Tools for functional dissection of site-specific O-GlcNAcylation. RSC Chem. Biol. 1, 98–109 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Stoevesandt, O. & Taussig, M. J. Phospho-specific antibodies by design. Nat. Biotechnol. 31, 889–891 (2013).

    Article  CAS  PubMed  Google Scholar 

  25. Alteen, M. G., Tan, H. Y. & Vocadlo, D. J. Monitoring and modulating O-GlcNAcylation: assays and inhibitors of O-GlcNAc processing enzymes. Curr. Opin. Struct. Biol. 68, 157–165 (2021).

    Article  CAS  PubMed  Google Scholar 

  26. Martin, S. E. S. et al. Structure-based evolution of low nanomolar O-GlcNAc transferase inhibitors. J. Am. Chem. Soc. 140, 13542–13545 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Gloster, T. M. et al. Hijacking a biosynthetic pathway yields a glycosyltransferase inhibitor within cells. Nat. Chem. Biol. 7, 174–181 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Ju Kim, E. O-GlcNAc transferase: structural characteristics, catalytic mechanism and small-molecule inhibitors. ChemBioChem 21, 3026–3035 (2020).

    Article  CAS  PubMed  Google Scholar 

  29. Liu, T. W. et al. Metabolic inhibitors of O-GlcNAc transferase that act in vivo implicate decreased O-GlcNAc levels in leptin-mediated nutrient sensing. Angew. Chem. Int. Ed. Engl. 57, 7644–7648 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Elbatrawy, A. A., Kim, E. J. & Nam, G. O-GlcNAcase: emerging mechanism, substrate recognition and small-molecule inhibitors. ChemMedChem 15, 1244–1257 (2020).

    Article  CAS  PubMed  Google Scholar 

  31. Selnick, H. G. et al. Discovery of MK-8719, a potent O-GlcNAcase inhibitor as a potential treatment for tauopathies. J. Med. Chem. 62, 10062–10097 (2019). This paper reports the medicinal chemistry efforts that led to the first FDA-sanctioned inhibitor of OGA in human disease.

    Article  CAS  PubMed  Google Scholar 

  32. Carrillo, L. D., Krishnamoorthy, L. & Mahal, L. K. A cellular FRET-based sensor for β-O-GlcNAc, a dynamic carbohydrate modification involved in signaling. J. Am. Chem. Soc. 128, 14768–14769 (2006).

    Article  CAS  PubMed  Google Scholar 

  33. Carrillo, L. D., Froemming, J. A. & Mahal, L. K. Targeted in vivo O-GlcNAc sensors reveal discrete compartment-specific dynamics during signal transduction. J. Biol. Chem. 286, 6650–6658 (2011).

    Article  CAS  PubMed  Google Scholar 

  34. Cecioni, S. & Vocadlo, D. J. Carbohydrate bis-acetal-based substrates as tunable fluorescence-quenched probes for monitoring exo-glycosidase activity. J. Am. Chem. Soc. 139, 8392–8395 (2017).

    Article  CAS  PubMed  Google Scholar 

  35. Lee, J.-H. et al. PET quantification of brain O-GlcNAcase with [18F]LSN3316612 in healthy human volunteers. EJNMMI Res. 10, 20 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Paul, S. et al. Evaluation of a PET radioligand to image O-GlcNAcase in brain and periphery of rhesus monkey and knock-out mouse. J. Nucl. Med. 60, 129–134 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Aguilar, A. L., Hou, X., Wen, L., Wang, P. G. & Wu, P. A chemoenzymatic histology method for O-GlcNAc detection. ChemBioChem 18, 2416–2421 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Haynes, P. A. & Aebersold, R. Simultaneous detection and identification of O-GlcNAc-modified glycoproteins using liquid chromatography−tandem mass spectrometry. Anal. Chem. 72, 5402–5410 (2000).

    Article  CAS  PubMed  Google Scholar 

  39. Vosseller, K. et al. Quantitative analysis of both protein expression and serine / threonine post-translational modifications through stable isotope labeling with dithiothreitol. Proteomics 5, 388–398 (2005).

    Article  CAS  PubMed  Google Scholar 

  40. Vosseller, K. et al. O-linked N-acetylglucosamine proteomics of postsynaptic density preparations using lectin weak affinity chromatography and mass spectrometry. Mol. Cell. Proteomics 5, 923–934 (2006).

    Article  CAS  PubMed  Google Scholar 

  41. Chalkley, R. J., Thalhammer, A., Schoepfer, R. & Burlingame, A. L. Identification of protein O-GlcNAcylation sites using electron transfer dissociation mass spectrometry on native peptides. Proc. Natl Acad. Sci. USA 106, 8894–8899 (2009).

    Article  PubMed Central  Google Scholar 

  42. Alfaro, J. F. et al. Tandem mass spectrometry identifies many mouse brain O-GlcNAcylated proteins including EGF domain-specific O-GlcNAc transferase targets. Proc. Natl Acad. Sci. USA 109, 7280–7285 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Darabedian, N. & Pratt, M. R. Identifying potentially O-GlcNAcylated proteins using metabolic labeling, bioorthogonal enrichment, and Western blotting. Methods Enzymol. 622, 293–307 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Qin, W. et al. Artificial cysteine S-glycosylation induced by per-O-acetylated unnatural monosaccharides during metabolic glycan labeling. Angew. Chem. Int. Ed. Engl. 57, 1817–1820 (2018). This paper reports off-target S-GlcNAcylatedartifactsin metabolic labeling, suggesting that stringent validation of O-GlcNAc proteomic studies is required.

    Article  CAS  PubMed  Google Scholar 

  45. Pedowitz, N. J. et al. Anomeric fatty acid functionalization prevents nonenzymatic S-glycosylation by monosaccharide metabolic chemical reporters. ACS Chem. Biol. https://doi.org/10.1021/acschembio.1c00470 (2021).

  46. Hao, Y. et al. Next-generation unnatural monosaccharides reveal that ESRRB O-GlcNAcylation regulates pluripotency of mouse embryonic stem cells. Nat. Commun. 10, 4065 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  47. Khidekel, N. et al. A chemoenzymatic approach toward the rapid and sensitive detection of O-GlcNAc posttranslational modifications. J. Am. Chem. Soc. 125, 16162–16163 (2003).

    Article  CAS  PubMed  Google Scholar 

  48. Woo, C. M. et al. Mapping and quantification of over 2000 O-linked glycopeptides in activated human T cells with isotope-targeted glycoproteomics (Isotag). Mol. Cell. Proteomics 17, 764–775 (2018). This paper reports exquisitely sensitive O-GlcNAc labeling tools with isotope tagging to enable global characterization of O-GlcNAc sites during T cell activation events.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Li, J. et al. An isotope-coded photocleavable probe for quantitative profiling of protein O-GlcNAcylation. ACS Chem. Biol. 14, 4–10 (2019).

    Article  CAS  PubMed  Google Scholar 

  50. Levine, P. M. et al. α-Synuclein O-GlcNAcylation alters aggregation and toxicity, revealing certain residues as potential inhibitors of Parkinson’s disease. Proc. Natl Acad. Sci. USA 116, 1511–1519 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Galesic, A. et al. Comparison of N-acetyl-glucosamine to other monosaccharides reveals structural differences for the inhibition of α-synuclein aggregation. ACS Chem. Biol. 16, 14–19 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Schwagerus, S., Reimann, O., Despres, C., Smet-Nocca, C. & Hackenberger, C. P. Semi-synthesis of a tag-free O-GlcNAcylated tau protein by sequential chemoselective ligation. J. Pept. Sci. 22, 327–333 (2016).

    Article  CAS  PubMed  Google Scholar 

  53. Lin, W., Gao, L. & Chen, X. Protein-specific imaging of O-GlcNAcylation in single cells. ChemBioChem 16, 2571–2575 (2015).

    Article  CAS  PubMed  Google Scholar 

  54. Ramirez, D. H. et al. Engineering a proximity-directed O-GlcNAc transferase for selective protein O-GlcNAcylation in cells. ACS Chem. Biol. 15, 1059–1066 (2020).

  55. Ge, Y. et al. Target protein deglycosylation in living cells by a nanobody-fused split O-GlcNAcase. Nat. Chem. Biol. 17, 593–600 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Boulard, M., Rucli, S., Edwards, J. R. & Bestor, T. H. Methylation-directed glycosylation of chromatin factors represses retrotransposon promoters. Proc. Natl Acad. Sci. USA 117, 14292–14298 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Adli, M. The CRISPR tool kit for genome editing and beyond. Nat. Commun. 9, 1911 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  58. Gorelik, A. et al. Genetic recoding to dissect the roles of site-specific protein O-GlcNAcylation. Nat. Struct. Mol. Biol. 26, 1071–1077 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Macauley, M. S., Stubbs, K. A. & Vocadlo, D. J. O-GlcNAcase catalyzes cleavage of thioglycosides without general acid catalysis. J. Am. Chem. Soc. 127, 17202–17203 (2005).

    Article  CAS  PubMed  Google Scholar 

  60. Kim, E. Y. et al. A role for O-GlcNAcylation in setting circadian clock speed. Genes Dev. 26, 490–502 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  61. Keembiyehetty, C. et al. Conditional knock-out reveals a requirement for O-linked N-acetylglucosaminase (O-GlcNAcase) in metabolic homeostasis. J. Biol. Chem. 290, 7097–7113 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  62. Okuyama, R. & Marshall, S. UDP-N-acetylglucosaminyl transferase (OGT) in brain tissue: temperature sensitivity and subcellular distribution of cytosolic and nuclear enzyme. J. Neurochem. 86, 1271–1280 (2003).

    Article  CAS  PubMed  Google Scholar 

  63. Levine, Z. G. et al. Mammalian cell proliferation requires noncatalytic functions of O-GlcNAc transferase. Proc. Natl Acad. Sci. USA 118, e2016778118 (2021). OGT has distinct protein-regulatory roles through three mechanisms: O-GlcNAc catalytic modification of proteins, O-GlcNAc-driven proteolysis, and non-catalytic functions. Rapid OGT regulatory tools are also developed herein.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  64. Konzman, D. et al. O-GlcNAc: regulator of signaling and epigenetics linked to X-linked intellectual disability. Front. Genet. 11, 605263 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  65. Pravata, V. M. et al. An intellectual disability syndrome with single-nucleotide variants in O-GlcNAc transferase. Eur. J. Hum. Genet 28, 706–714 (2020). X-linked intellectual disability is the first human disease implicated with OGT single-nucleotide polymorphisms, indicating a genetic basis (also see ref. 64).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Yuzwa, S. A. et al. A potent mechanism-inspired O-GlcNAcase inhibitor that blocks phosphorylation of tau in vivo. Nat. Chem. Biol. 4, 483–490 (2008).

    Article  CAS  PubMed  Google Scholar 

  67. Stewart, L. T. et al. Acute increases in protein O-GlcNAcylation dampen epileptiform activity in hippocampus. J. Neurosci. 37, 8207–8215 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  68. Lagerlöf, O. et al. The nutrient sensor OGT in PVN neurons regulates feeding. Science 351, 1293–1296 (2016). This paper reveals that organismal feeding behavior is controlled by O-GlcNAc, which is revealed to influence appetite at the molecular level in PVN neurons.

    Article  PubMed  PubMed Central  Google Scholar 

  69. Vaidyanathan, K. & Wells, L. Multiple tissue-specific roles for the O-GlcNAc post-translational modification in the induction of and complications arising from type II diabetes. J. Biol. Chem. 289, 34466–34471 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  70. Chen, P.-H. et al. Gigaxonin glycosylation regulates intermediate filament turnover and may impact giant axonal neuropathy etiology or treatment. JCI Insight 5, e127751 (2019).

  71. Abramowitz, L. K., Harly, C., Das, A., Bhandoola, A. & Hanover, J. A. Blocked O-GlcNAc cycling disrupts mouse hematopoeitic stem cell maintenance and early T cell development. Sci. Rep. 9, 12569 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  72. Baumann, D. et al. Role of nutrient-driven O-GlcNAc-post-translational modification in pancreatic exocrine and endocrine islet development. Development 147, dev186643 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  73. Hruby, A. & Hu, F. B. The epidemiology of obesity: a big picture. Pharmacoeconomics 33, 673–689 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  74. Wang, Z. et al. Extensive crosstalk between O-GlcNAcylation and phosphorylation regulates cytokinesis. Sci. Signal. 3, ra2 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

  75. Leney, A. C., El Atmioui, D., Wu, W., Ovaa, H. & Heck, A. J. R. Elucidating crosstalk mechanisms between phosphorylation and O-GlcNAcylation. Proc. Natl Acad. Sci. USA 114, E7255–E7261 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  76. Yang, X. et al. Phosphoinositide signalling links O-GlcNAc transferase to insulin resistance. Nature 451, 964–969 (2008).

    Article  CAS  PubMed  Google Scholar 

  77. Lambert, M., Bastide, B. & Cieniewski-Bernard, C. Involvement of O-GlcNAcylation in the skeletal muscle physiology and physiopathology: focus on muscle metabolism. Front. Endocrinol. (Lausanne) 9, 578 (2018).

    Article  Google Scholar 

  78. Yang, Y. et al. O-GlcNAc transferase inhibits visceral fat lipolysis and promotes diet-induced obesity. Nat. Commun. 11, 181 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  79. Parker, M. P., Peterson, K. R. & Slawson, C. O-GlcNAcylation and O-GlcNAc cycling regulate gene transcription: emerging roles in cancer. Cancers 13, 1666 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  80. Ozcan, S., Andrali, S. S. & Cantrell, J. E. Modulation of transcription factor function by O-GlcNAc modification. Biochim. Biophys. Acta 1799, 353–364 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

  81. Akella, N. M. et al. O-GlcNAc transferase regulates cancer stem-like potential of breast cancer cells. Mol. Cancer Res. 18, 585–598 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  82. Hrit, J. et al. OGT binds a conserved C-terminal domain of TET1 to regulate TET1 activity and function in development. eLife 7, e34870 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  83. Hornbeck, P. V. et al. PhosphoSitePlus, 2014: mutations, PTMs and recalibrations. Nucleic Acids Res. 43, D512–D520 (2015).

    Article  CAS  PubMed  Google Scholar 

  84. York, W. S. et al. GlyGen: computational and informatics resources for glycoscience. Glycobiology 30, 72–73 (2020).

    Article  CAS  PubMed  Google Scholar 

  85. Darabedian, N., Thompson, J. W., Chuh, K. N., Hsieh-Wilson, L. C. & Pratt, M. R. Optimization of chemoenzymatic mass tagging by strain-promoted cycloaddition (SPAAC) for the determination of O-GlcNAc stoichiometry by Western blotting. Biochemistry 57, 5769–5774 (2018).

    Article  CAS  PubMed  Google Scholar 

  86. Wells, L. et al. Mapping sites of <em>O</em>-GlcNAc modification using affinity tags for serine and threonine post-translational modifications. Mol. Cell. Proteomics 1, 791–804 (2002).

    Article  CAS  PubMed  Google Scholar 

  87. Gupta, R. & Brunak, S. Prediction of glycosylation across the human proteome and the correlation to protein function. Pac. Symp. Biocomput. 310–322 (2002).

  88. Jia, C., Zuo, Y. & Zou, Q. O-GlcNAcPRED-II: an integrated classification algorithm for identifying O-GlcNAcylation sites based on fuzzy undersampling and a K-means PCA oversampling technique. Bioinformatics 34, 2029–2036 (2018).

    Article  CAS  PubMed  Google Scholar 

  89. Jochmann, R., Holz, P., Sticht, H. & Sturzl, M. Validation of the reliability of computational O-GlcNAc prediction. Biochim. Biophys. Acta 1844, 416–421 (2014).

    Article  CAS  PubMed  Google Scholar 

  90. Anzalone, A. V. et al. Search-and-replace genome editing without double-strand breaks or donor DNA. Nature 576, 149–157 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  91. McKitrick, T. R. et al. Development of smart anti-glycan reagents using immunized lampreys. Commun. Biol. 3, 91 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  92. Zichel, R., Chearwae, W., Pandey, G. S., Golding, B. & Sauna, Z. E. Aptamers as a sensitive tool to detect subtle modifications in therapeutic proteins. PLoS One 7, e31948 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  93. Levine, Z. G. & Walker, S. The biochemistry of O-GlcNAc transferase: which functions make it essential in mammalian cells? Annu. Rev. Biochem. 85, 631–657 (2016).

    Article  CAS  PubMed  Google Scholar 

  94. Shen, D. L., Gloster, T. M., Yuzwa, S. A. & Vocadlo, D. J. Insights into O-linked N-acetylglucosamine ([0-9]O-GlcNAc) processing and dynamics through kinetic analysis of O-GlcNAc transferase and O-GlcNAcase activity on protein substrates. J. Biol. Chem. 287, 15395–15408 (2012). Detailed in vitro kinetic studies of OGT and OGA reveal mechanistic roles in nutrient sensing, adding to a rich body of OGT and OGA mechanistic studies.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  95. Levine, Z. G. et al. O-GlcNAc transferase recognizes protein substrates using an asparagine ladder in the tetratricopeptide repeat (TPR) superhelix. J. Am. Chem. Soc. 140, 3510–3513 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  96. Kositzke, A. et al. Elucidating the protein substrate recognition of O-GlcNAc transferase (OGT) toward O-GlcNAcase (OGA) using a GlcNAc electrophilic probe. Int. J. Biol. Macromol. 169, 51–59 (2021).

    Article  CAS  PubMed  Google Scholar 

  97. Toleman, C. A. et al. Structural basis of O-GlcNAc recognition by mammalian 14-3-3 proteins. Proc. Natl Acad. Sci. USA 115, 5956–5961 (2018). O-GlcNAcylation was long thought to prevent protein-protein interactions, but this report shows O-GlcNAc-driven interactions for the first time, confirmed by structural biology.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  98. Myers, S. A. et al. SOX2 O-GlcNAcylation alters its protein-protein interactions and genomic occupancy to modulate gene expression in pluripotent cells. eLife 5, e10647 (2016).

    Article  PubMed  PubMed Central  Google Scholar 

  99. Yu, S.-H. et al. Metabolic labeling enables selective photocrosslinking of O-GlcNAc-modified proteins to their binding partners. Proc. Natl Acad. Sci. USA 109, 4834–4839 (2012).

  100. Balana, A. T. et al. O-GlcNAc modification of small heat shock proteins enhances their anti-amyloid chaperone activity. Nat. Chem. 13, 441–450 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank P. Marino, Program Director at the US National Institute of General Medical Sciences (NIGMS) for her commitment and vision in expanding glycoscience as a national priority. We thank M. Bond and K. Krueger, Program Officers, for their support with glycobiology tools and for discussions and notes. We thank all participants of the international O-GlcNAc workshop, held in March 2020, who contributed to discussion, both during and after the meeting. We thank NIGMS for grant no. 1R35GM142637-01 (to C.F.) and the National Institutes of Diabetes and Digestive and Kidney Diseases for financial support (to J.A.H.).

Author information

Authors and Affiliations

Authors

Corresponding authors

Correspondence to Charlie Fehl or John A. Hanover.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Peer review information Nature Chemical Biology thanks Matthew Pratt, Xiaoyong Yang and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Fehl, C., Hanover, J.A. Tools, tactics and objectives to interrogate cellular roles of O-GlcNAc in disease. Nat Chem Biol 18, 8–17 (2022). https://doi.org/10.1038/s41589-021-00903-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41589-021-00903-6

Search

Quick links

Nature Briefing: Cancer

Sign up for the Nature Briefing: Cancer newsletter — what matters in cancer research, free to your inbox weekly.

Get what matters in cancer research, free to your inbox weekly. Sign up for Nature Briefing: Cancer