Qiu Y, Yu W, Zhang X, Zhang M, Ni Y, Lai S, Wu Q.
Upregulation of OGT-mediated EZH2 O-GlcNAcylation Promotes Human Umbilical Vein Endothelial Cell Proliferation, Invasion, Migration, and Tube Formation in Gestational Diabetes Mellitus.
Cell biochemistry and biophysics202539751742
Abstract: O-linked N-acetylglucosamine transferase (OGT)-catalyzed O-linked N-acetylglucosamine glycosylation (O-GlcNAcylation) is closely associated with diabetes progression. This study aims to investigate the mechanism of OGT in regulating endothelial dysfunction in gestational diabetes mellitus (GDM). Expressions of OGT, O-linked N-acetylglucosamine (O-GlcNAc), enhancer of zeste homolog 2 (EZH2), and HEK27me3 in human umbilical vein endothelial cells (HUVECs) and GDM-derived HUVECs (GDM-HUVECs) were assessed by western blot. RT-qPCR and western blot assays were used to test the OGT overexpression and EZH2 silencing levels. CCK-8, EdU, wound healing, and transwell invasion assays were used to analyze the cell proliferative, migratory, and invasive abilities. Tube formation assay was performed to evaluate angiogenesis ability of cells. Western blot assay was performed to estimate vascular endothelial growth factor (VEGF) and p-VEGFR2 levels in cells. The binding of O-GlcNAc and EZH2 after OGT overexpression was measured by Co-IP assay. The results showed that OGT, O-GlcNAc, EZH2, and HEK27me3 expressions were declined in GDM-HUVECs. OGT overexpression induced the proliferation, migration, and invasion of GDM-HUVECs, and also elevated angiogenesis and the expressions of VEGF and p-VEGFR2 in cells. O-GlcNAc, EZH2, and HEK27me3 expressions were upregulated after OGT overexpression. OGT upregulation induced the binding between O-GlcNAc and EZH2. EZH2 silencing attenuated the promotion of OGT overexpression on the proliferative, invasive, migratory, and angiogenic capacities of GDM-HUVECs. To be concluded, OGT overexpression stabilized EZH2 expression by promoting O-GlcNAcylation modification of EZH2, and further enhanced proliferation, migration, and invasion as well as angiogenesis of GDM-HUVECs. While these effects were decayed after EZH2 absenting. Overall, the modulation of OGT on endothelial dysfunction in GDM provides a novel perspective for the clinical treatment of GDM.
Hu M, Zhang X, Gao YP, Hu YX, Teng T, Wang SS, Tang QZ.
Isthmin-1 Improves Aging-Related Cardiac Dysfunction in Mice through Enhancing Glycolysis and SIRT1 Deacetylase Activity.
Aging and disease202438300636
Abstract: Aging-related cardiac dysfunction poses a major risk factor of mortality for elderly populations, however, efficient treatment for aging-related cardiac dysfunction is far from being known. Isthmin-1 (ISM1) is a novel adipokine that promotes glucose uptake and acts indispensable roles in restraining inflammatory and fibrosis. The present study aims to investigate the potential role and molecular mechanism of ISM1 in aging-related cardiac dysfunction. Aged and matched young mice were overexpressed or silenced with ISM1 to investigate the role of ISM1 in aging-related cardiac dysfunction. Moreover, H9C2 cells were stimulated with D-galactose (D-gal) to examine the role of ISM1 in vitro. Herein, we found that cardiac-specific overexpression of ISM1 significantly mitigated insulin resistance by promoting glucose uptake in aging mice. ISM1 overexpression alleviated while ISM1 silencing deteriorated cellular senescence, cardiac inflammation, and dysfunction in natural and accelerated cardiac aging. Mechanistically, ISM1 promoted glycolysis and activated Sirtuin-1 (SIRT1) through increasing glucose uptake. ISM1 increased glucose uptake via translocating GLUT4 to the surface, thereby enhancing glycolytic flux and hexosamine biosynthetic pathway (HBP) flux, ultimately leading to increased SIRT1 activity through O-GlcNAc modification. ISM1 may serve as a novel potential therapeutic target for preventing aging-related cardiac disease in elderly populations. ISM1 prevents aging-related cardiac dysfunction by promoting glycolysis and enhancing SIRT1 deacetylase activity, making it a promising therapeutic target for aging-related cardiac disease.
Hao Y, Li X, Qin K, Shi Y, He Y, Zhang C, Cheng B, Zhang X, Hu G, Liang S, Tang Q, Chen X.
Chemoproteomic and Transcriptomic Analysis Reveals that O-GlcNAc Regulates Mouse Embryonic Stem Cell Fate through the Pluripotency Network.
Angewandte Chemie (International ed. in English)2023
62(17)
36852467
Abstract: Self-renewal and differentiation of embryonic stem cells (ESCs) are influenced by protein O-linked β-N-acetylglucosamine (O-GlcNAc) modification, but the underlying mechanism remains incompletely understood. Herein, we report the identification of 979 O-GlcNAcylated proteins and 1340 modification sites in mouse ESCs (mESCs) by using a chemoproteomics method. In addition to OCT4 and SOX2, the third core pluripotency transcription factor (PTF) NANOG was found to be modified and functionally regulated by O-GlcNAc. Upon differentiation along the neuronal lineage, the O-GlcNAc stoichiometry at 123 sites of 83 proteins-several of which were PTFs-was found to decline. Transcriptomic profiling reveals 2456 differentially expressed genes responsive to OGT inhibition during differentiation, of which 901 are target genes of core PTFs. By acting on the core PTF network, suppression of O-GlcNAcylation upregulates neuron-related genes, thus contributing to mESC fate determination.
Zhang X, Li D, Zhu J, Zheng J, Li H, He Q, Peng J, Chen S, Chen XL, Wang W.
RNAPII Degradation Factor Def1 Is Required for Development, Stress Response, and Full Virulence of Magnaporthe oryzae.
Journal of fungi (Basel, Switzerland)2023
9(4)
37108921
Abstract: The RNA polymerase II degradation factor Degradation Factor 1 (Def1) is important for DNA damage repair and plays various roles in eukaryotes; however, the biological role in plant pathogenic fungi is still unknown. In this study, we investigated the role of Def1 during the development and infection of the rice blast fungus Magnaporthe oryzae. The deletion mutant of Def1 displayed slower mycelial growth, less conidial production, and abnormal conidial morphology. The appressoria of Δdef1 was impaired in the penetration into host cells, mainly due to blocking in the utilization of conidial storages, such as glycogen and lipid droplets. The invasive growth of the Δdef1 mutant was also retarded and accompanied with the accumulation of reactive oxygen species (ROS) inside the host cells. Furthermore, compared with the wild type, Δdef1 was more sensitive to multiple stresses, such as oxidative stress, high osmotic pressure, and alkaline/acidic pH. Interestingly, we found that Def1 was modified by O-GlcNAcylation at Ser232, which was required for the stability of Def1 and its function in pathogenicity. Taken together, the O-GlcNAc modified Def1 is required for hyphae growth, conidiation, pathogenicity, and stress response in M. oryzae. This study reveals a novel regulatory mechanism of O-GlcNAc-mediated Def1 in plant pathogenic fungi.
Su Y, Luo Y, Zhang P, Lin H, Pu W, Zhang H, Wang H, Hao Y, Xiao Y, Zhang X, Wei X, Nie S, Zhang K, Fu Q, Chen H, Huang N, Ren Y, Wu M, Chow BKC, Chen X, Jin W, Wang F, Zhao L, Rao F.
Glucose-induced CRL4(COP1)-p53 axis amplifies glycometabolism to drive tumorigenesis.
Molecular cell2023
83(13)
37390815
Abstract: The diabetes-cancer association remains underexplained. Here, we describe a glucose-signaling axis that reinforces glucose uptake and glycolysis to consolidate the Warburg effect and overcome tumor suppression. Specifically, glucose-dependent CK2 O-GlcNAcylation impedes its phosphorylation of CSN2, a modification required for the deneddylase CSN to sequester Cullin RING ligase 4 (CRL4). Glucose, therefore, elicits CSN-CRL4 dissociation to assemble the CRL4COP1 E3 ligase, which targets p53 to derepress glycolytic enzymes. A genetic or pharmacologic disruption of the O-GlcNAc-CK2-CSN2-CRL4COP1 axis abrogates glucose-induced p53 degradation and cancer cell proliferation. Diet-induced overnutrition upregulates the CRL4COP1-p53 axis to promote PyMT-induced mammary tumorigenesis in wild type but not in mammary-gland-specific p53 knockout mice. These effects of overnutrition are reversed by P28, an investigational peptide inhibitor of COP1-p53 interaction. Thus, glycometabolism self-amplifies via a glucose-induced post-translational modification cascade culminating in CRL4COP1-mediated p53 degradation. Such mutation-independent p53 checkpoint bypass may represent the carcinogenic origin and targetable vulnerability of hyperglycemia-driven cancer.
Yu F, Yang S, Ni H, Heng D, Wu X, Yang M, Zhang X, Cao Y, Pei Y, An D, Li D, Liu D, Liu L, Pan L, Chen Q, Zhu X, Zhou J.
O-GlcNAcylation Regulates Centrosome Behavior and Cell Polarity to Reduce Pulmonary Fibrosis and Maintain the Epithelial Phenotype.
Advanced science (Weinheim, Baden-Wurttemberg, Germany)2023
10(36)
37963851
Abstract: O-GlcNAcylation functions as a cellular nutrient and stress sensor and participates in almost all cellular processes. However, it remains unclear whether O-GlcNAcylation plays a role in the establishment and maintenance of cell polarity, because mice lacking O-GlcNAc transferase (OGT) are embryonically lethal. Here, a mild Ogt knockout mouse model is constructed and the important role of O-GlcNAcylation in establishing and maintaining cell polarity is demonstrated. Ogt knockout leads to severe pulmonary fibrosis and dramatically promotes epithelial-to-mesenchymal transition. Mechanistic studies reveal that OGT interacts with pericentriolar material 1 (PCM1) and centrosomal protein 131 (CEP131), components of centriolar satellites required for anchoring microtubules to the centrosome. These data further show that O-GlcNAcylation of PCM1 and CEP131 promotes their centrosomal localization through phase separation. Decrease in O-GlcNAcylation prevents PCM1 and CEP131 from localizing to the centrosome, instead dispersing these proteins throughout the cell and impairing the microtubule-centrosome interaction to disrupt centrosome positioning and cell polarity. These findings identify a previously unrecognized role for protein O-GlcNAcylation in establishing and maintaining cell polarity with important implications for the pathogenesis of pulmonary fibrosis.
Zhang J, Yang P, Liu D, Gao M, Wang J, Yu T, Zhang X, Liu Y.
Inhibiting Hyper-O-GlcNAcylation of c-Myc accelerate diabetic wound healing by alleviating keratinocyte dysfunction.
Burns & trauma2021
9
34646892
Abstract: Diabetic foot ulcers characterized by delayed healing are one of the main complications of diabetes. Epidermal keratinocyte dysfunction has been found to play a pivotal role in the poor healing ability of diabetic wounds. In this study, we aimed to explore the relationship between c-Myc and its O-linked N-acetylglucosamine (O-GlcNAc) glycosylation (O-GlcNAcylation) modification and keratinocyte dysfunction in diabetic wounds.
Chen LL, Huang JQ, Wu YY, Chen LB, Li SP, Zhang X, Wu S, Ren FZ, Lei XG.
Loss of Selenov predisposes mice to extra fat accumulation and attenuated energy expenditure.
Redox biology2021
45
34167027
Abstract: Selenoprotein V (SELENOV) is a new and the least conserved member of the selenoprotein family. Herein we generated Selenov knockout (KO) mice to determine its in vivo function. The KO led to 16-19% increases (P < 0.05) in body weight that were largely due to 54% higher (P < 0.05) fat mass accumulation, compared with the wild-type (WT) controls. The extra fat accumulation in the KO mice was mediated by up-regulations of genes and proteins involved in lipogenesis (Acc, Fas, Dgat, and Lpl; up by 40%-1.1-fold) and down-regulations of lipolysis (Atgl, Hsl, Ces1d, and Cpt1a; down by 36-89%) in the adipose tissues. The KO also decreased (P < 0.05) VO2 consumption (14-21%), VCO2 production (14-16%), and energy expenditure (14-23%), compared with the WT controls. SELENOV and O-GlcNAc transferase (OGT) exhibited a novel protein-protein interaction that explained the KO-induced decreases (P < 0.05) of OGT protein (15-29%), activity (33%), and function (O-GlcNAcylation, 10-21%) in the adipose tissues. A potential cascade of SELENOV-OGT-AMP-activated protein kinase might serve as a central mechanism to link the biochemical and molecular responses to the KO. Overall, our data revealed a novel in vivo function and mechanism of SELENOV as a new inhibitor of body fat accumulation, activator of energy expenditure, regulator of O-GlcNAcylation, and therapeutic target of such related disorders.
Zhu G, Qian M, Lu L, Chen Y, Zhang X, Wu Q, Liu Y, Bian Z, Yang Y, Guo S, Wang J, Pan Q, Sun F.
O-GlcNAcylation of YY1 stimulates tumorigenesis in colorectal cancer cells by targeting SLC22A15 and AANAT.
Carcinogenesis2019
40(9)
30715269
Abstract: Emerging studies have revealed that O-GlcNAcylation plays pivotal roles in the tumorigenesis of colorectal cancers (CRCs). However, the underlying mechanism still remains largely unknown. Here, we demonstrated that Yin Yang 1 (YY1) was O-GlcNAcylated by O-GlcNAc transferase (OGT) and O-GlcNAcylation of YY1 could increase the protein expression by enhancing its stability. O-GlcNAcylation facilitated transformative phenotypes of CRC cell in a YY1-dependent manner. Also, O-GlcNAcylation stimulates YY1-dependent transcriptional activity. Besides, we also identified the oncoproteins, SLC22A15 and AANAT, which were regulated by YY1 directly, are responsible for the YY1 stimulated tumorigenesis. Furthermore, we identified the main putative O-GlcNAc site of YY1 at Thr236, and mutating of this site decreased the pro-tumorigenic capacities of YY1. We concluded that O-GlcNAcylation of YY1 stimulates tumorigenesis in CRC cells by targeting SLC22A15 and AANAT, suggesting that YY1 O-GlcNAcylation might be a potential effective therapeutic target for treating CRC.
Chen Y, Zhu G, Liu Y, Wu Q, Zhang X, Bian Z, Zhang Y, Pan Q, Sun F.
O-GlcNAcylated c-Jun antagonizes ferroptosis via inhibiting GSH synthesis in liver cancer.
Cellular signalling2019
63
31394193
Abstract: Ferroptosis is a metabolism-related cell death. Stimulating ferroptosis in liver cancer cells is a strategy to treat liver cancer. However, how to eradicate liver cancer cells through ferroptosis and the obstacles to inducing ferroptosis in liver cancer remain unclear. Here, we observed that erastin suppressed the malignant phenotypes of liver cancer cells by inhibiting O-GlcNAcylation of c-Jun and further inhibited protein expression, transcription activity and nuclear accumulation of c-Jun. Overexpression of c-Jun-WT with simultaneous PuGNAc treatment conversely inhibited erastin-induced ferroptosis, whereas overexpression of c-Jun-WT alone or overexpression of c-Jun-S73A (a non-O-GlcNAcylated form of c-Jun) with PuGNAc treatment did not exert a similar effect. GSH downregulation induced by erastin was restored by overexpression of c-Jun-WT with simultaneous PuGNAc treatment. In addition, overexpression of c-Jun-WT, but not its S73A mutant, induced PSAT1 and CBS transcription via directly binding to their promoter regions, suggesting that GSH synthesis is regulated by O-GlcNAcylated c-Jun. A positive correlation between c-Jun O-GlcNAcylation and GSH was observed in clinical samples. Collectively, O-GlcNAcylated c-Jun represents an obstructive factor to ferroptosis, and targeting O-GlcNAcylated c-Jun might be helpful for treating liver cancer.
Song N, Qi Q, Cao R, Qin B, Wang B, Wang Y, Zhao L, Li W, Du X, Liu F, Yan Y, Yi W, Jiang H, Li T, Zhou T, Li HY, Xia Q, Zhang XM, Zhong W, Li AL, Duan X.
MAVS O-GlcNAcylation Is Essential for Host Antiviral Immunity against Lethal RNA Viruses.
Cell reports2019
28(9)
31461653
Abstract: It is known that lethal viruses profoundly manipulate host metabolism, but how the metabolism alternation affects the immediate host antiviral immunity remains elusive. Here, we report that the O-GlcNAcylation of mitochondrial antiviral-signaling protein (MAVS), a key mediator of interferon signaling, is a critical regulation to activate the host innate immunity against RNA viruses. We show that O-GlcNAcylation depletion in myeloid cells renders the host more susceptible to virus infection both in vitro and in vivo. Mechanistically, we demonstrate that MAVS O-GlcNAcylation is required for virus-induced MAVS K63-linked ubiquitination, thereby facilitating IRF3 activation and IFNβ production. We further demonstrate that D-glucosamine, a commonly used dietary supplement, effectively protects mice against a range of lethal RNA viruses, including human influenza virus. Our study highlights a critical role of O-GlcNAcylation in regulating host antiviral immunity and validates D-glucosamine as a potential therapeutic for virus infections.
Zhang X, Shu XE, Qian SB.
O-GlcNAc modification of eIF4GI acts as a translational switch in heat shock response.
Nature chemical biology2018
14(10)
30127386
Abstract: Heat shock response (HSR) is an ancient signaling pathway leading to thermoprotection of nearly all living organisms. Emerging evidence suggests that intracellular O-linked β-N-acetylglucosamine (O-GlcNAc) serves as a molecular 'thermometer' by reporting ambient temperature fluctuations. Whether and how O-GlcNAc modification regulates HSR remains unclear. Here we report that, upon heat shock stress, the key translation initiation factor eIF4GI undergoes dynamic O-GlcNAcylation at the N-terminal region. Without O-GlcNAc modification, the preferential translation of stress mRNAs is impaired. Unexpectedly, stress mRNAs are entrapped within stress granules (SGs) that are no longer dissolved during stress recovery. Mechanistically, we show that stress-induced eIF4GI O-GlcNAcylation repels poly(A)-binding protein 1 and promotes SG disassembly, thereby licensing stress mRNAs for selective translation. Using various eIF4GI mutants created by CRISPR/Cas9, we demonstrate that eIF4GI acts as a translational switch via reversible O-GlcNAcylation. Our study reveals a central mechanism linking heat stress sensing, protein remodeling, SG dynamics and translational reprogramming.
Zhang X, Wang L, Lak B, Li J, Jokitalo E, Wang Y.
GRASP55 Senses Glucose Deprivation through O-GlcNAcylation to Promote Autophagosome-Lysosome Fusion.
Developmental cell2018
45(2)
29689198
Abstract: The Golgi apparatus is the central hub for protein trafficking and glycosylation in the secretory pathway. However, how the Golgi responds to glucose deprivation is so far unknown. Here, we report that GRASP55, the Golgi stacking protein located in medial- and trans-Golgi cisternae, is O-GlcNAcylated by the O-GlcNAc transferase OGT under growth conditions. Glucose deprivation reduces GRASP55 O-GlcNAcylation. De-O-GlcNAcylated GRASP55 forms puncta outside of the Golgi area, which co-localize with autophagosomes and late endosomes/lysosomes. GRASP55 depletion reduces autophagic flux and results in autophagosome accumulation, while expression of an O-GlcNAcylation-deficient mutant of GRASP55 accelerates autophagic flux. Biochemically, GRASP55 interacts with LC3-II on the autophagosomes and LAMP2 on late endosomes/lysosomes and functions as a bridge between LC3-II and LAMP2 for autophagosome and lysosome fusion; this function is negatively regulated by GRASP55 O-GlcNAcylation. Therefore, GRASP55 senses glucose levels through O-GlcNAcylation and acts as a tether to facilitate autophagosome maturation.
Chen Y, Liu R, Chu Z, Le B, Zeng H, Zhang X, Wu Q, Zhu G, Chen Y, Liu Y, Sun F, Lu Z, Qiao Y, Wang J.
High glucose stimulates proliferative capacity of liver cancer cells possibly via O-GlcNAcylation-dependent transcriptional regulation of GJC1.
Journal of cellular physiology2018
234(1)
30078215
Abstract: Although it is generally accepted that diabetes is one of the most important risk factors for liver cancer, the underlying mechanism is still not well understood. The purpose of the current study is to further investigate how high concentrations of glucose (HG), a major symptom of diabetes, stimulate the development of liver malignancy. Using data mining, gap junction protein gamma 1 (GJC1) was identified as a critical proto-oncoprotein that is essential for the HG stimulation of proliferative capacity in liver cancer cells. Furthermore, enhanced transcriptional expression of GJC1 might occur after stimulation by HG. A transcription factor zinc finger protein 410 (APA1)-binding motif was found to be located at the -82 to -77 nt region within the GJC1 promoter. Without APA1, HG was unable to increase GJC1 expression. Interestingly, APA1, but not GJC1, can be O-GlcNAcylated in liver cancer cells. Moreover, O-GlcNAcylation is essential for HG-induced APA1 binding to the GJC1 promoter. Notably, global O-GlcNAcylation and expression of APA1 and GJC1 were highly elevated in liver cancer patients with diabetes compared to those in patients without diabetes. The HG-stimulated proliferative capacity was abolished upon decreasing O-GlcNAcylation, which could be reversed gradually by the simultaneous overexpression of APA1 and GJC1. Therefore, GJC1 could be a potential target for preventing liver cancer in patients with diabetes.
Zhang X, Qiao Y, Wu Q, Chen Y, Zou S, Liu X, Zhu G, Zhao Y, Chen Y, Yu Y, Pan Q, Wang J, Sun F.
The essential role of YAP O-GlcNAcylation in high-glucose-stimulated liver tumorigenesis.
Nature communications2017
8
28474680
Abstract: O-GlcNAcylation has been implicated in the tumorigenesis of various tissue origins, but its function in liver tumorigenesis is not clear. Here, we demonstrate that O-GlcNAcylation can enhance the expression, stability and function of Yes-associated protein (YAP), the downstream transcriptional regulator of the Hippo pathway and a potent oncogenic factor in liver cancer. O-GlcNAcylation induces transformative phenotypes of liver cancer cells in a YAP-dependent manner. An O-GlcNAc site of YAP was identified at Thr241, and mutating this site decreased the O-GlcNAcylation, stability, and pro-tumorigenic capacities of YAP, while increasing YAP phosphorylation. Importantly, we found via in vitro cell-based and in vivo mouse model experiments that O-GlcNAcylation of YAP was required for high-glucose-induced liver tumorigenesis. Interestingly, a positive feedback between YAP and global cellular O-GlcNAcylation is also uncovered. We conclude that YAP O-GlcNAcylation is a potential therapeutic intervention point for treating liver cancer associated with high blood glucose levels and possibly diabetes.
Han C, Gu Y, Shan H, Mi W, Sun J, Shi M, Zhang X, Lu X, Han F, Gong Q, Yu W.
O-GlcNAcylation of SIRT1 enhances its deacetylase activity and promotes cytoprotection under stress.
Nature communications2017
8(1)
29133780
Abstract: SIRT1 is the most evolutionarily conserved mammalian sirtuin, and it plays a vital role in the regulation of metabolism, stress responses, genome stability, and ageing. As a stress sensor, SIRT1 deacetylase activity is significantly increased during stresses, but the molecular mechanisms are not yet fully clear. Here, we show that SIRT1 is dynamically modified with O-GlcNAc at Ser 549 in its carboxy-terminal region, which directly increases its deacetylase activity both in vitro and in vivo. The O-GlcNAcylation of SIRT1 is elevated during genotoxic, oxidative, and metabolic stress stimuli in cellular and mouse models, thereby increasing SIRT1 deacetylase activity and protecting cells from stress-induced apoptosis. Our findings demonstrate a new mechanism for the activation of SIRT1 under stress conditions and suggest a novel potential therapeutic target for preventing age-related diseases and extending healthspan.
Wang Y, Liu J, Jin X, Zhang D, Li D, Hao F, Feng Y, Gu S, Meng F, Tian M, Zheng Y, Xin L, Zhang X, Han X, Aravind L, Wei M.
O-GlcNAcylation destabilizes the active tetrameric PKM2 to promote the Warburg effect.
Proceedings of the National Academy of Sciences of the United States of America2017
114(52)
29229835
Abstract: The Warburg effect, characterized by increased glucose uptake and lactate production, is a well-known universal across cancer cells and other proliferating cells. PKM2, a splice isoform of the pyruvate kinase (PK) specifically expressed in these cells, serves as a major regulator of this metabolic reprogramming with an adjustable activity subjected to numerous allosteric effectors and posttranslational modifications. Here, we have identified a posttranslational modification on PKM2, O-GlcNAcylation, which specifically targets Thr405 and Ser406, residues of the region encoded by the alternatively spliced exon 10 in cancer cells. We show that PKM2 O-GlcNAcylation is up-regulated in various types of human tumor cells and patient tumor tissues. The modification destabilized the active tetrameric PKM2, reduced PK activity, and led to nuclear translocation of PKM2. We also observed that the modification was associated with an increased glucose consumption and lactate production and enhanced level of lipid and DNA synthesis, indicating that O-GlcNAcylation promotes the Warburg effect. In vivo experiments showed that blocking PKM2 O-GlcNAcylation attenuated tumor growth. Thus, we demonstrate that O-GlcNAcylation is a regulatory mechanism for PKM2 in cancer cells and serves as a bridge between PKM2 and metabolic reprogramming typical of the Warburg effect.
Qiao Y, Zhang X, Zhang Y, Wang Y, Xu Y, Liu X, Sun F, Wang J.
High Glucose Stimulates Tumorigenesis in Hepatocellular Carcinoma Cells Through AGER-Dependent O-GlcNAcylation of c-Jun.
Diabetes2016
65(3)
26825459
Abstract: Epidemiologic studies suggest that hepatocellular carcinoma (HCC) has a strong relationship with diabetes. However, the underlying molecular mechanisms still remain unclear. Here, we demonstrated that high glucose (HG), one of the main characteristics of diabetes, was capable of accelerating tumorigenesis in HCC cells. Advanced glycosylation end product-specific receptor (AGER) was identified as a stimulator during this process. Mechanistically, AGER activated a hexosamine biosynthetic pathway, leading to enhanced O-GlcNAcylation of target proteins. Notably, AGER was capable of increasing activity and stability of proto-oncoprotein c-Jun via O-GlcNAcylation of this protein at Ser73. Interestingly, c-Jun can conversely enhance AGER transcription. Thereby, a positive autoregulatory feedback loop that stimulates diabetic HCC was established. Finally, we found that AG490, an inhibitor of Janus kinase, has the ability to impair AGER expression and its functions in HCC cells. In conclusion, AGER and its functions to stimulate O-GlcNAcylation are important during liver tumorigenesis, when high blood glucose levels are inadequately controlled.
Yao B, Xu Y, Wang J, Qiao Y, Zhang Y, Zhang X, Chen Y, Wu Q, Zhao Y, Zhu G, Sun F, Li Z, Yuan H.
Reciprocal regulation between O-GlcNAcylation and tribbles pseudokinase 2 (TRIB2) maintains transformative phenotypes in liver cancer cells.
Cellular signalling2016
28(11)
27515988
Abstract: TRIB2 has been identified as an onco-protein, and O-GlcNAcylation of target proteins has been reported to stimulate transformative phenotypes in liver cancer cells. However, the relationships between TRIB2 and O-GlcNAcylation are still unknown. The aim of this study was to investigate whether and how O-GlcNAcylation and TRIB2 regulate each other. We found that stimulation of O-GlcNAcylation elevates TRIB2 by enhancing its protein stability. TRIB2 can be O-GlcNAcylated by the hexosamine biosynthesis pathway (HBP). Also, O-GlcNAcylation boosting of transformative phenotypes of liver cancer cells might occur in a TRIB2-dependent manner. Interestingly, TRIB2 stimulated the metabolism of HBP, demonstrating that TRIB2 has positive feedback on O-GlcNAcylation. Notably, TRIB2 was found to maintain the stability of guanylate cyclase 1 alpha 3 (GUCY1A3), a key component of HBP, by interacting GUCY1A3 and reducing its ubiquitination. Importantly, TRIB2-dependent regulation of metabolism, transformative phenotypes, and O-GlcNAcylation all rely on GUCY1A3. Mouse experiments demonstrate that O-GlcNAcylation of TRIB2 is much higher in the livers of diabetic mice compared to control mice, suggesting that O-GlcNAcylation of TRIB2 might be critical for diabetes-associated liver cancer. Collectively, we have uncovered a positive auto-regulatory feedback between O-GlcNAcylation and TRIB2, which might be regarded as a promising therapeutic target for liver cancer.
Han C, Shan H, Bi C, Zhang X, Qi J, Zhang B, Gu Y, Yu W.
A highly effective and adjustable dual plasmid system for O-GlcNAcylated recombinant protein production in E. coli.
Journal of biochemistry2015
157(6)
25619971
Abstract: O-GlcNAcylation is a ubiquitous, dynamic and reversible post-translational protein modification in metazoans, and it is catalysed and removed by O-GlcNAc transferase (OGT) and O-GlcNAcase, respectively. Prokaryotes lack endogenous OGT activity. It has been reported that coexpression of mammalian OGT with its target substrates in Escherichia coli produce O-GlcNAcylated recombinant proteins, but the plasmids used were not compatible, and the expression of both OGT and its target protein were induced by the same inducer. Here, we describe a compatible dual plasmid system for coexpression of OGT and its target substrate for O-GlcNAcylated protein production in E. coli. The approach was validated using the CKII and p53 protein as control. This compatible dual plasmid system contains an arabinose-inducible OGT expression vector with a pUC origin and an isopropyl β-d-thiogalactopyranoside-inducible OGT target substrate expression vector bearing a p15A origin. The dual plasmid system produces recombinant proteins with varying O-GlcNAcylation levels by altering the inducer concentration. More importantly, the O-GlcNAcylation efficiency was much higher than the previously reported system. Altogether, we established an adjustable compatible dual plasmid system that can effectively yield O-GlcNAcylated proteins in E. coli.
Zhang X, Bennett V.
Identification of O-linked N-acetylglucosamine modification of ankyrinG isoforms targeted to nodes of Ranvier.
The Journal of biological chemistry1996
271(49)
8940148
Abstract: AnkyrinGs of 270 and 480 kDa are localized at nodes of Ranvier and are candidates to couple the voltage-dependent sodium channel and neurofascin to the spectrin/actin network. This study presents evidence that these ankyrins contain O-linked GlcNAc residues and identifies as the site of glycosylation a serine-rich domain that distinguishes them from other ankyrin isoforms. The 480-kDa ankyrinG, extracted from brain membranes associated with wheat germ agglutinin-affinity columns, was [3H]galactose-labeled with UDP-[3H] galactose and galactosyltransferase, and cross-reacted with an antibody against O-GlcNAc monosaccharides. AnkyrinG-associated sugars are O-linked monosaccharides based on resistance to peptide-N-glycosidase F and analysis of saccharides released by beta-elimination. The serine-rich domain is the site of glycosylation based on wheat germ agglutinin binding activity of polypeptides produced by in vitro translation in reticulocyte lysates. Immunofluorescence revealed co-localization of ankyrinG and O-GlcNAc immunoreactivity at nodes of Ranvier. These observations suggest that ankyrin at the node of Ranvier is O-GlcNAc-glycosylated and are the first demonstration of a post-translational modification that is concentrated at the node of Ranvier and not in adjacent areas of myelinated axons.