Project description:Single Cell RNAseq was performed to identify cell types and upstream regulators in pancreatic epithelial cells with or without OGT.
Project description:To gain comprehensive insight into the OGT-dependent transcriptional program in Treg cells, we performed RNA-sequencing of isolated YFP+ Treg cells from Foxp3YFP-Cre/wtOgtwt/fl and healthy Foxp3YFP-Cre/wtOgtfl/fl females to avoid secondary changes in gene expression caused by inflammation. We were able to identify 269 differentially expressed genes including 154 downregulated and 115 upregulated with p values less than 0.01, OGT-deficient Treg cells had impaired suppressive function and attenuated IL2/STAT5 signaling pathway.
Project description:Over a billion people suffer from chronic liver diseases worldwide, which often leads to fibrosis and then cirrhosis. Treatments for fibrosis remain experimental, in part because no unifying mechanism has been identified that initiates liver fibrosis. O-linked β-N-acetylglucosamine (O-GlcNAc) transferase (OGT) plays a pro-survival role under stress in many tissues. Here we report that OGT protects against hepatocyte necroptosis and initiation of liver fibrosis. Decreased O-GlcNAc levels were seen in patients with alcoholic liver cirrhosis and in mice with ethanol-induced liver injury. Liver-specific O-GlcNAc transferase (OGT) knockout (OGT-LKO) mice progressed to liver fibrosis at 10 weeks of age. OGT-deficient hepatocytes underwent necroptosis. These findings identify OGT as a key suppressor of hepatocyte necroptosis and OGT-LKO mice may serve as an effective spontaneous genetic model of liver fibrosis.
Project description:To investigate the roles of Ogt in smooth muscle cells (SMCs), we crossed Ogt floxed mice and SM22-CreER mice to generate inducible SMC Ogt specific knockout mice. Aorta tissues from these mice were isolated and RNA were extracted and performed RNAseq analysis.
Project description:TET2 directly interacts with OGT, which is important for the chromatin association of OGT in vivo. Although this specific interaction does not regulate the enzymatic activity of TET2, it facilitates OGT-dependent histone O-GlcNAcylation. Moreover, OGT associates with TET2 at transcription starting sites (TSS). Down-regulation of TET2 reduces the amount of H2B S112 GlcNAc marks in vivo, which are associated with gene transcription regulation. We found that OGT interacts with TET2 tightly. Using ChIP-seq with specific antibodies, we tested the co-localization of TET2 and OGT in genome level.
Project description:Ogt catalyzed O-linked N-acetylglucosamine (O-GlcNAcylation, O-GlcNAc) plays an important function in diverse biological processes and diseases. However, the roles of Ogt in regulating neurogenesis remain largely unknown. Here, we show that Ogt deficiency or depletion in adult neural stem cells (aNSCs) leads to the diminishment of aNSCs pool and the aberrant neurogenesis, and consequently impairs cognitive function of adult mice. RNA sequencing reveals that Ogt deficiency alters the transcription of genes relating to cell cycle, neurogenesis, and neuronal development. Mechanistic studies show that Ogt directly interacts with Notch1 and catalyzes the O-GlcNAc modification of Notch TM/ICD fragment. The decreased O-GlcNAc modification of TM/ICD increases the binding of E3 ubiquitin ligase Itch to TM/ICD and promotes its degradation. Itch knockdown rescues neurogenic defects in Ogt-deficient mice. OGT also regulates human cortical neurogenesis in forebrain organoids derived from induced pluripotent stem cells. Our findings reveal the essential roles and mechanisms of Ogt and O-GlcNAc modification in regulating mammalian neurogenesis and cognition.
Project description:O-linked N-acetylglucosamine (O-GlcNAc ) transferase (OGT) activity is essential for embryonic stem (ES) cell viability and mouse development. OGT is present in both cytoplasm and nucleus of different cell types and mediates serine or threonine glycosylation. The Ogt gene locus resides on the X-chromosome and its activity is required for the viability of male ES cells. Using Ogt conditional knock out (KO) ES cells it was shown the failure of establishing stable KO ES clones further suggesting that Ogt activity is required for ES cell self-renewal and pluripotency. For understanding these changes, we performed global gene expression upon silencing of Ogt mediated by esiRNA in mouse Embryonic Stem Cells. Total RNA was extracted from E14 ES cells (derived from Ola129 mice) transiently transfected with Ogt specific and control esiRNA oligos 48 hrs. post-transfection.