Detection and identification of <i>O</i>‐GlcNAcylated proteins by proteomic approaches

Vercoutter‐Edouart, Anne‐Sophie; Yazidi‐Belkoura, Ikram El; Guinez, Céline; Baldini, Steffi F.; Leturcq, Maïté; Mortuaire, Marlène; Mir, Anne-Marie; Steenackers, Agata; Dehennaut, Vanessa; Pierce, Annick; Lefebvre, Tony

doi:10.1002/pmic.201400326

Cited by 32 publications

(26 citation statements)

References 96 publications

(185 reference statements)

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“…Loss of O-GlcNAcylation was shown to result in loss of cellular function or even cell death [45] and the disturbance of normal O-GlcNAc function has been linked to many diseases, including Alzheimer's disease, diabetes, and other chronic illnesses [46,47]. Only a single OGT enzyme is responsible for O-GlcNAcylation of over a thousand protein substrates [48]. Only a single OGT enzyme is responsible for O-GlcNAcylation of over a thousand protein substrates [48].…”

Section: Regulation Of O-glcnacylationmentioning

confidence: 99%

“…Therefore, the proper functioning of O-GlcNAcylation in cells is essential, however, how this regulation is fine-tuned is still not well understood. Only a single OGT enzyme is responsible for O-GlcNAcylation of over a thousand protein substrates [48]. OGT has three isoforms that differ in the number of tetratricopeptide repeats (TPRs) they harbor at their N terminus, a well-described protein-protein interaction-mediating domain [49].…”

Section: Regulation Of O-glcnacylationmentioning

confidence: 99%

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Crosstalk between phosphorylation and O‐GlcNAcylation: friend or foe

2018

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A wide variety of protein post-translational modifications (PTMs) decorate cellular proteins, regulating their structure, interactions and ultimately their function. The density of co-occurring PTMs on proteins can be very high, where multiple PTMs can positively or negatively influence each other's actions, termed PTM crosstalk. In this review, we highlight recent progress in the area of PTM crosstalk, whereby we focus on crosstalk between protein phosphorylation and O-GlcNAcylation. These two PTMs largely target identical (i.e., Ser and Thr) amino acids in proteins. Phosphorylation/O-GlcNAcylation crosstalk comes in many flavors, for instance by competition for the same site/residue (reciprocal crosstalk), as well as by modifications influencing each other in proximity or even distal on the protein sequence. PTM crosstalk is observed on the writers of these modifications (i.e., kinases and O-GlcNAc transferase), on the erasers (i.e., phosphatases and O-GlcNAcase), and on the readers and the substrates. We describe examples of all these different flavors of crosstalk, and additionally the methods that are emerging to better investigate in particular phosphorylation/O-GlcNAcylation crosstalk.

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Section: Regulation Of O-glcnacylationmentioning

confidence: 99%

Section: Regulation Of O-glcnacylationmentioning

confidence: 99%

Crosstalk between phosphorylation and O‐GlcNAcylation: friend or foe

2018

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“…Most of the proteins are subjected to covalent chemical modifications that occur co- or post-translationally. Post-translational modifications (PTMs) are very diverse and consist in the addition—usually enzymatically—of simple or complex (e.g., peptides, proteins, glycosylphosphatidylinositol) groups, or in the proteolytic cleavage to enlarge the complexity of the proteome ( 1 ). To date, many hundreds of PTMs have been described among which are found a wide variety of glycosylations including the O -linked β-N-acetylglucosaminylation ( O -GlcNAcylation).…”

Section: Introductionmentioning

confidence: 99%

Apart From Rhoptries, Identification of Toxoplasma gondii's O-GlcNAcylated Proteins Reinforces the Universality of the O-GlcNAcome

et al. 2018

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O-linked β-N-acetylglucosaminylation or O-GlcNAcylation is a widespread post-translational modification that belongs to the large and heterogeneous group of glycosylations. The functions managed by O-GlcNAcylation are diverse and include regulation of transcription, replication, protein's fate, trafficking, and signaling. More and more evidences tend to show that deregulations in the homeostasis of O-GlcNAcylation are involved in the etiology of metabolic diseases, cancers and neuropathologies. O-GlcNAc transferase or OGT is the enzyme that transfers the N-acetylglucosamine residue onto target proteins confined within the cytosolic and nuclear compartments. A form of OGT was predicted for Toxoplasma and recently we were the first to show evidence of O-GlcNAcylation in the apicomplexans Toxoplasma gondii and Plasmodium falciparum. Numerous studies have explored the O-GlcNAcome in a wide variety of biological models but very few focus on protists. In the present work, we used enrichment on sWGA-beads and immunopurification to identify putative O-GlcNAcylated proteins in Toxoplasma gondii. Many of the proteins found to be O-GlcNAcylated were originally described in higher eukaryotes and participate in cell shape organization, response to stress, protein synthesis and metabolism. In a more original way, our proteomic analyses, confirmed by sWGA-enrichment and click-chemistry, revealed that rhoptries, proteins necessary for invasion, are glycosylated. Together, these data show that regardless of proteins strictly specific to organisms, O-GlcNAcylated proteins are rather similar among living beings.

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“…In addition to antibody-based detection techniques of global O -GlcNAc-pattern identification, recently more and more studies emerged, investigating into sequence-specific localization of functional alterations caused by protein modification with O -GlcNAc. While these methods certainly will provide an important leap in understanding complex biological regulatory circuits, the analytical techniques, mainly relying on mass spectrometry combined with soft ionization methods, are still more suitable for focusing on an individual candidate protein than on global effects [ 44 , 54 ]. Nevertheless, future studies will have to integrate these techniques for more detailed description of senescence-associated changes in specific O -GlcNAcylation.…”

Section: Discussionmentioning

confidence: 99%

Senescence-Associated Changes in Proteome andO-GlcNAcylation Pattern in Human Peritoneal Mesothelial Cells

Tarantino

Rudolf

Aufricht

et al. 2015

BioMed Research International

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Introduction. Senescence of peritoneal mesothelial cells represents a biological program defined by arrested cell growth and altered cell secretory phenotype with potential impact in peritoneal dialysis. This study aims to characterize cellular senescence at the level of global protein expression profiles and modification of proteins with O-linked N-acetylglucosamine (O-GlcNAcylation). Methods. A comparative proteomics analysis between young and senescent human peritoneal mesothelial cells (HPMC) was performed using two-dimensional gel electrophoresis. O-GlcNAc status was assessed by Western blot under normal conditions and after modulation with 6-diazo-5-oxo-L-norleucine (DON) to decrease O-GlcNAcylation or O-(2-acetamido-2-deoxy-D-glucopyranosylidene) amino N-phenyl carbamate (PUGNAc) to increase O-GlcNAcylation. Results. Comparison of protein pattern of senescent and young HPMC revealed 29 differentially abundant protein spots, 11 of which were identified to be actin (cytoplasmic 1 and 2), cytokeratin-7, cofilin-2, transgelin-2, Hsp60, Hsc70, proteasome β-subunits (type-2 and type-3), nucleoside diphosphate kinase A, and cytosolic 5′(3′)-deoxyribonucleotidase. Although the global level of O-GlcNAcylation was comparable, senescent cells were not sensitive to modulation by PUGNAc. Discussion. This study identified changes of the proteome and altered dynamics of O-GlcNAc regulation in senescent mesothelial cells. Whereas changes in cytoskeleton-associated proteins likely reflect altered cell morphology, changes in chaperoning and housekeeping proteins may have functional impact on cellular stress response in peritoneal dialysis.

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Detection and identification of O‐GlcNAcylated proteins by proteomic approaches

Cited by 32 publications

References 96 publications

Crosstalk between phosphorylation and O‐GlcNAcylation: friend or foe

Crosstalk between phosphorylation and O‐GlcNAcylation: friend or foe

Apart From Rhoptries, Identification of Toxoplasma gondii's O-GlcNAcylated Proteins Reinforces the Universality of the O-GlcNAcome

Senescence-Associated Changes in Proteome andO-GlcNAcylation Pattern in Human Peritoneal Mesothelial Cells

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