O-GlcNAcylation in mitochondrial protein in rat astrocytes
Ontology highlight
ABSTRACT:
INSTRUMENT(S):
ORGANISM(S): Rattus Norvegicus (rat)
TISSUE(S): Brain, Fibroblast
DISEASE(S): Disease Free
SUBMITTER: 
Kazuhide Hayakawa
LAB HEAD: Kazuhide Hayakawa
PROVIDER: PXD045316 | Pride | 2023-10-20
REPOSITORIES: Pride
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| 48685_47167.mzXML | Mzxml | |||
| 48685_73965.mzid.gz | Mzid | |||
| 48685_73965.pep.xml | Pepxml | |||
| 48685_out.tgz | Other | |||
| 48685_sequest.params | Other |
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Project description:O-linked-N-acetylglucosamine (O-GlcNAc) has emerged as a critical regulator of diverse cellular processes, but its role in embryonic stem cells (ESCs) and pluripotency has not been investigated. Here we show that O-GlcNAcylation directly regulates core components of the pluripotency network. Blocking O-GlcNAcylation disrupts ESC self-renewal and reprogramming of somatic cells to induced pluripotent stem cells. The core reprogramming factors Oct4 and Sox2 are O-GlcNAcylated in ESCs, but the O-GlcNAc modification is rapidly removed upon differentiation. O-GlcNAc modification of Threonine 228 in Oct4 regulates Oct4 transcriptional activity and is important for inducing many pluripotency related genes, including Klf2, Klf5, Nr5a2, Tbx3 and Tcl1. A T228A point mutation that eliminates this O-GlcNAc modification reduces the capacity of Oct4 to maintain ESC self-renewal and reprogram somatic cells. Overall, our study makes a direct connection between O-GlcNAcylation of key regulatory transcription factors and the activity of the pluripotency network. 2 of E14 stem cell, 2 of embryonic body at day 2, 2 of embryonic body at day 2 treated with streptozotocin, 1 of ZHBTc4 stem cell treated with doxicyclin, and 1 of ZHBTc4 stem cell treated with doxicyclin and streptozotocin were analysed
2012-07-05 | E-GEOD-36322 | biostudies-arrayexpress
Project description:O-linked-N-acetylglucosamine (O-GlcNAc) has emerged as a critical regulator of diverse cellular processes, but its role in embryonic stem cells (ESCs) and pluripotency has not been investigated. Here we show that O-GlcNAcylation directly regulates core components of the pluripotency network. Blocking O-GlcNAcylation disrupts ESC self-renewal and reprogramming of somatic cells to induced pluripotent stem cells. The core reprogramming factors Oct4 and Sox2 are O-GlcNAcylated in ESCs, but the O-GlcNAc modification is rapidly removed upon differentiation. O-GlcNAc modification of Threonine 228 in Oct4 regulates Oct4 transcriptional activity and is important for inducing many pluripotency related genes, including Klf2, Klf5, Nr5a2, Tbx3 and Tcl1. A T228A point mutation that eliminates this O-GlcNAc modification reduces the capacity of Oct4 to maintain ESC self-renewal and reprogram somatic cells. Overall, our study makes a direct connection between O-GlcNAcylation of key regulatory transcription factors and the activity of the pluripotency network.
2012-07-06 | GSE36322 | GEO
Project description:Nutrient-driven O-GlcNAcylation of key components of the transcription machinery may epigenetically modulate gene expression in metazoans. Knockouts of the O-GlcNAc cycling enzymes in C. elegans are viable and fertile, allowing a global analysis of the impact of GlcNAcylation. Whole genome ChIP profiling of the O-GlcNAc cycling mutants identified over 800 genes associated with GlcNAcylated chromatin. This novel chromatin mark is preferentially found with genes involved in stress, longevity, and innate immunity, the same set of processes affected by global changes in gene expression analysis. This experiment collected data on fed animals (3 hours) for comparisons to previoulsy collected starved data to see what acute effects nutritional flux would have on either O-GlcNAc chromatin marks or RNA Pol II distributions.
2010-01-01 | GSE18611 | GEO
Project description:O-linked N-acetylglucosamination (O-GlcNAcylation) of proteins is increasingly recognized as an important cellular regulatory mechanism. In the heart, nucleocytoplasmic O-GlcNAcylation is involved in the modulation of multiple pathophysiological processes. However, the mechanisms leading to O-GlcNAcylation in mitochondria and the consequences on their function remain poorly understood. In this study, we used an in vitro reconstitution assay to characterize the intra-mitochondrial O-GlcNAc cycling system without potential confounding effects of O-GlcNAcylation in other cellular compartments. We performed a comparative analysis of the O-GlcNAcylome of isolated cardiac mitochondria acutely (30 min) exposed to UDP-GlcNAc in presence/absence of NButGT, a specific O-GlcNAcylation inducer, and evaluated the impact on mitochondrial function. 412 O-GlcNAcylated mitochondrial proteins were identified by mass spectrometry analysis, with 189 (Q<0.05) displaying increased O-GlcNAcylation in response to NButGT. Among the O-GlcNAcylated pathways identified, oxidative phosphorylation was the main affected. These acute changes in protein O-GlcNAcylation were associated with enhanced Complex I (CI) activity, increased maximal respiration in presence of CI substrates, and a striking reduction of mitochondrial ROS release, which could be related to O-GlcNAcylation of subunits within the NADH dehydrogenase module of CI. In conclusion, our work underlines the existence of a dynamic mitochondrial O-GlcNAcylation system capable of rapidly modifying mitochondrial function.
2022-02-11 | PXD026495 | Pride
Project description:O-GlcNAcylation is a reversible and dynamic post-translational protein modification catalyzed by O-GlcNAc transferase (OGT). Despite the reported association of O-GlcNAcylation with cancer metastasis, the O-GlcNAc proteome profile for cancer aggressiveness remains largely uncharac-terized. Here we report our comparative O-GlcNAc proteome profiling of 2 differentially invasive lung adenocarcinoma cell lines, which identified 158 down-regulated and 106 up-regulated can-didates in highly invasive cells. Among these differential proteins, a nuclear RNA-binding protein SAM68 (SRC associated in mitosis of 68 kDa) was further investigated. Results showed that SAM68 is O-GlcNAcylated and may interact with OGT in the nucleus. Eleven O-GlcNAcylation sites were identified, and data from mutant analysis suggested that multiple serine residues in the N-terminal region are important for O-GlcNAcylation and the function of SAM68 in modulating cancer cell migration and invasion. Analysis of clinical specimens found that high SAM68 ex-pression was associated with late cancer stages, and patients with high-OGT/high-SAM68 ex-pression in their tumors had poorer overall survival compared to those with low-OGT/low-SAM68 expression. Our study has revealed an invasiveness-associated O-GlcNAc proteome profile and connected O-GlcNAcylated SAM68 to lung cancer aggressiveness.
2022-02-17 | PXD029627 | Pride
Project description:O-linked β-N-acetylglucosaminylation (O-GlcNAcylation) is a ubiquitous posttranslational modification occurring in both animals and plants. Thousands of proteins along with their O-GlcNAcylation sites have been identified in various animal systems, yet the O-GlcNAcylated proteomes in plants remain poorly understood. Here, we report a large-scale profiling of protein O-GlcNAcylation in a site-specific manner in rice. We first established the metabolic glycan labeling strategy (MGL) with N-azidoacetylgalactosamine (GalNAz) in rice seedlings, which enabled incorporation of azides as a bioorthogonal handle into O-GlcNAc. By conjugation of the azide-incorporated O-GlcNAc with alkyne-biotin containing a cleavable linker via click chemistry, O-GlcNAcylated proteins were selectively enriched for mass spectrometry (MS) analysis. A total of 1,591 unambiguous O-GlcNAcylation sites distributed on 709 O-GlcNAcylated proteins were identified. Additionally, 102 O-GlcNAcylated proteins were identified with their O-GlcNAcylation sites located within a serine/threonine-enriched peptide, causing ambiguous site assignment. The identified O-GlcNAcylated proteins are involved in multiple biological processes such as transcription, translation and plant hormone signaling. Furthermore, we discovered two O-GlcNAc transferases (OsOGTs) in rice. By expressing OsOGTs in Escherichia coli and Nicotiana benthamiana leaves, we confirmed their OGT enzymatic activities and used them to validate the identified rice O-GlcNAcylated proteins. Our dataset provides a valuable resource for studying O-GlcNAc biology in rice, and the MGL method should facilitate the identification of O-GlcNAcylated proteins in various plants.
2023-05-10 | PXD036527 | Pride
Project description:We have found that histone H2B is GlcNAcylated at residue S112 by O-GlcNAc transferase and that H2B S112 GlcNAcylation fluctuates in response to extracellular glucose level. We have also found that H2B S112 GlcAcylation promotes H2B K120 ubiquitination. To investigate whether these histone modification correlate to transcriptional activation, we performed comprehensive gene expression analysis using Affymetrix GeneChip in HeLa cell cultured with different conditions, i.e. without glucose, with glucose and with FBS.
2011-10-27 | GSE33050 | GEO
Project description:Dysfunctional mitochondria and generation of reactive oxygen species (ROS) promote chronic diseases, which have spurred interest in the molecular mechanisms underlying these conditions. Previously, we have demonstrated that disruption of post-translational modification of proteins with β-linked N-acetylglucosamine (O- glcnAcylation) via overexpression of the O-glcnAc–regulating enzymes O- glcnAc transferase (OGT) or O- glcnAcase (OGA) impairs mitochondrial function. Here, we report that sustained alterations in O- glcnAcylation either by pharmacological or genetic manipulation also alters metabolic function. Sustained O-glcnAc elevation in SH-SY5Y neuroblastoma cells increased OGA expression and reduced cellular respiration and ROS generation. Cells with elevated O-glcnAc levels had elongated mitochondria and increased mitochondrial membrane potential, and RNA-Seq in SH-SY5Y cells indicated transcriptome reprogramming and down regulation of the NRF2-mediated antioxidant response. Sustained O-glcnAcylation in mice brain and liver validated the metabolic phenotypes observed in the cells, and OGT knockdown in the liver elevated ROS levels, impaired respiration, and increased the NRF2 antioxidant response. Moreover, elevated O-glcnAc levels promoted weight loss and lowered respiration in mice and skewed the mice toward carbohydrate-dependent metabolism as determined by indirect calorimetry. In summary, sustained elevation in O-glcnAcylation coupled with increased OGA expression reprograms energy metabolism, a finding that has potential implications for the etiology, development, and management of metabolic diseases.
2017-07-20 | GSE101642 | GEO
Project description:We have found that histone H2B is GlcNAcylated at residue S112 by O-GlcNAc transferase and that H2B S112 GlcNAcylation fluctuates in response to extracellular glucose level. We have also found that H2B S112 GlcAcylation promotes H2B K120 ubiquitination. To investigate whether these histone modification correlate to transcriptional activation, we performed comprehensive gene expression analysis using Affymetrix GeneChip in HeLa cell cultured with different conditions, i.e. without glucose, with glucose and with FBS. HeLa cells were cultured in DMEM with the following three conditions, 1) DMEM without glucose for 24 hours, 2) DMEM without glucose for 24 hours and followed by treatment with 4.5 g/L glucose for another 24 hours, 3) normal culture condition (DMEM with FBS). Total RNA was purified from these cells and each RNA was linearly amplified and hybridized to Affymetrix GeneChip.
2011-10-26 | E-GEOD-33050 | biostudies-arrayexpress
Project description:O-GlcNAcylation occurs on thousands of proteins involved in various cellular events. O-GlcNAc transferase (OGT), one of the enzymes responsible for protein O-GlcNAcylation, is known to be auto-O-GlcNAcylated at multiple sites. However, the role of O-GlcNAcylation on OGT is still unknown. Here, we report the role of O-GlcNAcylation on short form OGT (sOGT). By LC-ETD-MS analysis and western blot, we show that sOGT was mainly O-GlcNAcylated in the tetratricopeptide repeat (TPR) motifs with T12 and S56 being two “key” sites. T12 was a predominant O-GlcNAcylation site and the absence of S56 O-GlcNAcylation enhanced sOGT O-GlcNAcylation level. We found that O-GlcNAcylation of sOGT did not affect the enzyme activity but increased its binding to substrate proteins. S56A bound to and hence glycosylated more proteins, while T12A associated with fewer substrates. Proteomic studies combined with cell proliferation/cell cycle analyses indicated that S56A substantially enhanced cell proliferation, while T12A reduced it, and T12A led to a G2/M cell cycle arrest, due to O-GlcNAcylation of diverse proteins. These findings demonstrate that the O-GlcNAc pattern on sOGT modulates its function in cells by targeting different proteins.
2019-09-16 | PXD010173 | Pride