O-GlcNAcylation regulates c-Myc/PDK2 axis to rewire TCA cycle and promote colorectal tumor growth
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE232258 | GEO | 2024/05/10
REPOSITORIES: GEO
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Project description:TET proteins convert 5-methylcytosine to 5-hydroxymethylcytosine, an emerging dynamic epigenetic state of DNA that can influence transcription. Evidence has linked TET1 function to epigenetic repression complexes, yet mechanistic information, especially for the TET2 and TET3 proteins, remains limited. Here, we show a direct interaction of TET2 and TET3 with O-GlcNAc transferase (OGT). OGT does not appear to influence hmC activity, rather TET2 and TET3 promote OGT activity. TET2/3-OGT co-localize on chromatin at active promoters enriched for H3K4me3 and reduction of either TET2/3 or OGT activity results in a direct decrease in H3K4me3 and concomitant decreased transcription. Further, we show that Host Cell Factor 1 (HCF1), a component of the H3K4 methyltransferase SET1/COMPASS complex, is a specific GlcNAcylation target of TET2/3-OGT, and modification of HCF1 is important for the integrity of SET1/COMPASS. Additionally, we find both TET proteins and OGT activity promote binding of the SET1/COMPASS H3K4 methyltransferase, SETD1A, to chromatin. Finally, studies in Tet2 knockout mouse bone marrow tissue extend and support the data as decreases are observed of global GlcNAcylation and also of H3K4me3, notably at several key regulators of haematopoiesis. Together, our results unveil a step-wise model, involving TET-OGT interactions, promotion of GlcNAcylation, and influence on H3K4me3 via SET1/COMPASS, highlighting a novel means by which TETs may induce transcriptional activation. ChIP-Seq experiments were performed on Illumina HiScanSQ sequencer in wild-type HEK293T cells for H3K4me3 histone marks, O-GlcNAc and HCF1, for HT-TET2, HT-TET3 and HT-OGT in HEK293T cells overexpressing those three fusion proteins and in TET2 Kd HEK293T cells for H3K4me3 histone marks. ChIP-Seqs were also performed in mouse bone marrow tissues for H3K4me3 histone marks, O-GlcNAc, endogenous Tet2 and in Tet2 Ko bone marrow tissues for H3K4me3 histone marks.
2013-02-07 | E-GEOD-36620 | biostudies-arrayexpress
Project description:Many cancer cells display enhanced glycolysis and suppressed mitochondrial metabolism. This phenomenon, known as the Warburg effect, is critical for tumor development. However, how cancer cells coordinate glucose metabolism through glycolysis and the mitochondrial tricarboxylic acid (TCA) cycle is largely unknown. We demonstrate here that phosphoglycerate kinase 1 (PGK1), the first ATP-producing enzyme in glycolysis, is reversibly and dynamically modified with O-linked N-acetylglucosamine (O-GlcNAc) at threonine 255 (T255). O-GlcNAcylation activates PGK1 activity to enhance lactate production, and simultaneously induces PGK1 translocation into mitochondria. Inside mitochondria, PGK1 acts as a kinase to inhibit pyruvate dehydrogenase (PDH) complex to reduce oxidative phosphorylation. Blocking T255 O-GlcNAcylation of PGK1 decreases colon cancer cell proliferation, suppresses glycolysis, enhances the TCA cycle, and inhibits tumor growth in xenograft models. Furthermore, PGK1 O-GlcNAcylation levels are elevated in human colon cancers. This study highlights O-GlcNAcylation as an important signal for coordinating glycolysis and the TCA cycle to promote tumorigenesis.
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Project description:Despite the pivotal role of MYC in the pathogenesis of T-cell acute lymphoblastic leukemia (T-ALL) and many other cancers, the mechanisms underlying MYC-mediated tumorigenesis remain inadequately understood. Here we utilized a well-characterized zebrafish model of Myc-induced T-ALL for genetic studies to identify novel genes contributing to disease onset. We found that heterozygous inactivation of a tricarboxylic acid (TCA) cycle enzyme, dihydrolipoamide S-succinyltransferase (Dlst), significantly delayed tumor onset in zebrafish without detectable effects on fish development. DLST is the E2 transferase of the ?-ketoglutarate (?-KG) dehydrogenase complex (KGDHC), which converts ?-KG to succinyl-CoA in the TCA cycle. RNAi knockdown of DLST led to decreased cell viability and induction of apoptosis in human T-ALL cell lines. Polar metabolomics profiling revealed that the TCA cycle was disrupted by DLST knockdown in human T-ALL cells, as demonstrated by an accumulation of ?-KG and a decrease of succinyl-CoA. Addition of succinate, the downstream TCA cycle intermediate, to human T-ALL cells was sufficient to rescue defects in cell viability caused by DLST inactivation. Together, our studies uncovered an important role for DLST in MYC-mediated leukemogenesis and demonstrated the metabolic dependence of T-lymphoblasts on the TCA cycle, thus providing implications for targeted therapy.
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Project description:Mounting evidence points to alterations in mitochondrial metabolism in renal cell carcinoma (RCC). However, the mechanisms that regulate the TCA cycle in RCC remain uncharacterized. Here, we demonstrate that loss of TCA cycle enzyme expression is retained in RCC metastatic tissues. Moreover, proteomic analysis demonstrates that reduced TCA cycle enzyme expression is far more pronounced in RCC relative to other tumor types. Loss of TCA cycle enzyme expression is correlated with reduced expression of the transcription factor PGC-1α, which is also lost in RCC tissues. PGC-1α reexpression in RCC cells restores the expression of TCA cycle enzymes in vitro and in vivo and leads to enhanced glucose carbon incorporation into TCA cycle intermediates. Mechanistically, TGF-β signaling, in concert with histone deacetylase 7 (HDAC7), suppresses TCA cycle enzyme expression. Our studies show that pharmacologic inhibition of TGF-β restores the expression of TCA cycle enzymes and suppresses tumor growth in an orthotopic model of RCC. Taken together, this investigation reveals a potentially novel role for the TGF-β/HDAC7 axis in global suppression of TCA cycle enzymes in RCC and provides insight into the molecular basis of altered mitochondrial metabolism in this malignancy.
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Project description:OGT (O-GlcNAc transferase) is the distinctive enzyme responsible for catalyzing O-GlcNAc to the serine or threonine residues of thousands of cytoplasm and nuclear proteins that are involved in DNA damage, RNA splicing, and transcription preinitiation and initiation complex assembly. However, the molecular mechanism by OGT regulating gene transcription remains elusive. Using proximity labeling based mass spectrometry, we searched for functional partners of OGT and found that Dot1L, the conserved and unique histone methyltransferase mediated histone H3 lys79 methylation required for gene transcription, DNA damage repair, cell proliferation, and embryo development, interacts with OGT. Although this specific interaction does not regulate the enzymatic activity of Dot1L, it facilitates OGT-dependent histones O-GlcNAcylation. Moreover, OGT associates with Dot1L at transcription start sites, and depleting Dot1L decreased OGT associated with chromatin globally. Notably, downregulation of Dot1L reduces the levels of histone H2B S112 O-GlcNAcylation and histone H2B K120 ubiquitination in vivo, which are associated with gene transcription regulation. Taken together, these results reveal a Dot1L-dependent O-GlcNAcylation of chromatin.
2022-04-06 | GSE200059 | GEO
Project description:Drosophila development is a complex and dynamic process regulated, in part, by members of the Polycomb (Pc), Trithorax (Trx) and Compass chromatin modifier complexes. O-GlcNAc Transferase (OGT/SXC) is essential for Pc repression suggesting that the O-GlcNAcylation of proteins plays a key role in regulating development. OGT transfers N-acetyl-D-glucosamine (GlcNAc) onto hydroxyl groups of serine or threonine residues of key transcriptional regulators using the nutrient-derived UDP-GlcNAc as a substrate, which is dynamically removed by O-GlcNAcase (OGA). We performed ChIP-chip and microarray analysis after OGT or OGA RNAi knockdown in Drosophila S2 cells and found that O-GlcNAc was elevated genome wide particularly at genes related to mitosis and cell cycle in OGA RNAi cells, but not at sites co-occupied by Pc member Pleiohomeotic (Pho), such as the Hox and NK homeobox gene clusters. Microarray analysis suggested that altered O-GlcNAc cycling perturbed the expression of genes associated with morphogenesis and cell cycle regulation. To examine the in vivo consequences of disturbed O-GlcNAc cycling in the whole animal, we produced a null allele of oga (ogadel.1) in Drosophila. Epigenetic activators including Trx group members Trithorax (Trx), Absent small or homeotic discs 1 (Ash1) and Compass member Set1 histone methyltransferases are O-GlcNAc modified in ogadel.1 mutants. ogadel.1 mutants displayed altered expression of a distinct set of cell cycle related genes in ovaries. Our results suggest that the loss of OGA could affect epigenetic machinery by accumulating O-GlcNAc on numerous chromatin factors including Trx, Ash1 and Set1 in Drosophila. We performed affymetrix tilingarray analysis after OGT or OGA RNAi knockdown in Drosophila S2 cells to find if that O-GlcNAc was elevated genome wide particularly at genes related to mitosis and cell cycle in OGA RNAi cells, but not at sites co-occupied by Pc member Pleiohomeotic (Pho), ------------------------------- This represents the gene expression component only
2016-07-21 | E-GEOD-74846 | biostudies-arrayexpress
Project description:O-GlcNAc transferase (OGT) is overexpressed in aggressive prostate cancer. Here, we employed ChIP-seq to map chromatin-bound O-GlcNAc loci in prostate cancer cells and discovered that these overlap with sites of active transcription and MYC binding. Using RNA-seq, we show that inhibition of OGT promotes MYC-dependent transcriptional repression of mRNAs involved in G1-S transition. O-GlcNAc ChIP-seq regions are highly enriched to transcription start sites and identify the ‘GFY’-motif. Proteins binding to this motif have not been established and we use synthetic oligonucleotides as a bait to enrich protein complexes associated with this sequence. By comparing the unbiased proteomic data from oligonucleotide enrichment with proteomic data from O-GlcNAc and MYC ChIP-mass spectrometry, we identified host cell factor 1 (HCF-1) as an interaction partner of MYC. Inhibition of OGT disrupted the interaction between MYC and HCF-1, and compromised MYC’s ability to promote proliferation of prostate cancer cells in the absence of androgens. Reverse phase protein arrays identified a set of proteins involved in mitosis that are dependent on MYC and OGT activity for expression. In conclusion, we show that OGT activity regulates MYC-driven proliferation by coordinating transcription and translation of cell cycle genes.
2019-05-21 | GSE121472 | GEO
Project description:O-GlcNAc transferase (OGT) is overexpressed in aggressive prostate cancer. Here, we employed ChIP-seq to map chromatin-bound O-GlcNAc loci in prostate cancer cells and discovered that these overlap with sites of active transcription and MYC binding. Using RNA-seq, we show that inhibition of OGT promotes MYC-dependent transcriptional repression of mRNAs involved in G1-S transition. O-GlcNAc ChIP-seq regions are highly enriched to transcription start sites and identify the ‘GFY’-motif. Proteins binding to this motif have not been established and we use synthetic oligonucleotides as a bait to enrich protein complexes associated with this sequence. By comparing the unbiased proteomic data from oligonucleotide enrichment with proteomic data from O-GlcNAc and MYC ChIP-mass spectrometry, we identified host cell factor 1 (HCF-1) as an interaction partner of MYC. Inhibition of OGT disrupted the interaction between MYC and HCF-1, and compromised MYC’s ability to promote proliferation of prostate cancer cells in the absence of androgens. Reverse phase protein arrays identified a set of proteins involved in mitosis that are dependent on MYC and OGT activity for expression. In conclusion, we show that OGT activity regulates MYC-driven proliferation by coordinating transcription and translation of cell cycle genes.
2019-05-21 | GSE112667 | GEO
Project description:Heart failure is a leading cause of death worldwide, and failing heart muscle is marked by increased O-GlcNAcylation (OGN). It is unknown if excessive OGN contributes to cardiomyopathy and heart failure. OGN modifies serines and threonines, total OGN levels follow cellular nutrient and metabolic flux in addition to net activity of O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA). We developed new transgenic mouse models with myocardial delimited over-expression of OGA and OGT, and found that OGT transgenic mice developed severe cardiomyopathy and premature death. In contrast, OGA transgenic hearts had normal function, but were resistant to pathological stress. Interbreeding OGA transgenic mice rescued cardiomyopathy and premature death in OGT transgenic mice. RNA Seq and functional studies highlighted disrupted metabolism in hearts from OGT transgenic mice that was rescued by OGA transgenic interbreeding. Here we show excessive OGN causes cardiomyopathy, identify gene programs responsive to pathological OGN, and suggest attenuation of OGN may be an effective therapy for heart failure.
2021-07-10 | GSE134121 | GEO
Project description:Cancers exhibit altered metabolism characterized by increased glucose and glutamine uptake. The hexosamine biosynthetic pathway (HBP) uses glucose and glutamine, and directly contributes to O-linked-?-N-acetylglucosamine (O-GlcNAc) modifications on intracellular proteins. Multiple tumor types contain elevated total O-GlcNAcylation, in part, by increasing O-GlcNAc transferase (OGT) levels, the enzyme that catalyzes this modification. Although cancer cells require OGT for oncogenesis, it is not clear how tumor cells regulate OGT expression and O-GlcNAcylation. Here, it is shown that the PI3K-mTOR-MYC signaling pathway is required for elevation of OGT and O-GlcNAcylation in breast cancer cells. Treatment with PI3K and mTOR inhibitors reduced OGT protein expression and decreased levels of overall O-GlcNAcylation. In addition, both AKT and mTOR activation is sufficient to elevate OGT/O-GlcNAcylation. Downstream of mTOR, the oncogenic transcription factor c-MYC is required and sufficient for increased OGT protein expression in an RNA-independent manner and c-MYC regulation of OGT mechanistically requires the expression of c-MYC transcriptional target HSP90A. Finally, mammary tumor epithelial cells derived from MMTV-c-myc transgenic mice contain elevated OGT and O-GlcNAcylation and OGT inhibition in this model induces apoptosis. Thus, OGT and O-GlcNAcylation levels are elevated via activation of an mTOR/MYC cascade.Evidence indicates OGT as a therapeutic target in c-MYC-amplified cancers.
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