OGT binds a conserved C-terminal domain of TET1 to regulate TET1 activity and function in development

Abstract

TET enzymes convert 5-methylcytosine to 5-hydroxymethylcytosine and higher oxidized derivatives. TETs stably associate with and are post-translationally modified by the nutrient-sensing enzyme OGT, suggesting a connection between metabolism and the epigenome. Here, we show for the first time that modification by OGT enhances TET1 activity in vitro. We identify a TET1 domain that is necessary and sufficient for binding to OGT and report a point mutation that disrupts the TET1-OGT interaction. We show that this interaction is necessary for TET1 to rescue hematopoetic stem cell production in tet mutant zebrafish embryos, suggesting that OGT promotes TET1’s function during development. Finally, we show that disrupting the TET1-OGT interaction in mouse embryonic stem cells changes the abundance of TET2 and 5-methylcytosine, which is accompanied by alterations in gene expression. These results link metabolism and epigenetic control, which may be relevant to the developmental and disease processes regulated by these two enzymes.

Unraveling the functional role of DNA methylation using targeted DNA demethylation by steric blockage of DNA methyltransferase with CRISPR/dCas9

“…1J), clearly indicating a demethylation-independent transactivation capacity. It is indeed known that even the restricted catalytic TET domains used in dCas9-TET fusions retain a protein interaction domain that binds O-linked N-acetylglucosamine transferase (OGT) [28,29] and TET proteins and OGT have been shown to co-localize across the genome [98]. The recruited OGT regulates gene expression by glycosylating and modulating the activity of transcription factors such as HCFC1, SP1, OCT4, MYC, p53, and RNA polymerase II as well as histones to directly increase local H2B mono-ubiquitination and trimethylation of histone 3 on lysine 4, both of which are associated with increased gene expression [28,98,99].…”

“…More recently, the TET dioxygenases -which oxidize the methyl moiety in cytosine and can lead to passive loss of methylation by either inhibiting methylation during replication or through repair of the oxidized methylcytosine and its replacement with an unmethylated cytosine -were targeted to specific sites using a fusion of TET dioxygenase domains to catalytically inactive CRISPR/Cas9 (deltaCas9 or dCas9) [17][18][19][20]. However, this method still introduces several confounding factors that preclude causational inferences, such as the fact that oxidized methylcytosines are new epigenetic modifications that are not unmethylated cytosines [21][22][23][24][25][26][27] and the fact that TET has methylation-independent transcriptional activation activity [28,29]. We propose and optimize instead an enzyme-free CRISPR/dCas9-based system for targeted methylation editing which we show to be able to achieve selective methylation in vitro and passive demethylation in cells through steric interference with DNA methyltransferase activity.…”

O-GlcNAc: Regulator of Signaling and Epigenetics Linked to X-linked Intellectual Disability

“…Further, O-GlcNAcylation of TET1 was shown to regulate TET1 stability (Shi et al, 2013a), and OGT enhanced TET1 activity in vitro (Hrit et al, 2018). Disrupting the OGT-TET1 interaction in mouse ESCs resulted in increased 5mC and compensatory increases in TET2, accompanied by transcriptional changes (Hrit et al, 2018). These data indicate that the TET1-OGT complex is critical for proper pluripotency gene regulatory networks which maintain stem cell identity.…”

“…This interaction was required for OGT binding to chromatin and enhanced TET1 activity (Vella et al, 2013). Further, O-GlcNAcylation of TET1 was shown to regulate TET1 stability (Shi et al, 2013a), and OGT enhanced TET1 activity in vitro (Hrit et al, 2018). Disrupting the OGT-TET1 interaction in mouse ESCs resulted in increased 5mC and compensatory increases in TET2, accompanied by transcriptional changes (Hrit et al, 2018).…”

Epigenetic Dysregulation at the Crossroad of Women’s Cancer

“…Functions of several cancer promoting proteins, including, MYC and p53, are positively regulated by their O-linked-β-N-acetylglucosamine (O-GlcNAc) modification by O-GlcNAc transferase (OGT) enzyme [72]. The activity of TET1 in system development is positively regulated by its interaction with OGT [73]. Although OGT is known to be upregulated in breast cancer [74], its relationship with TET1 function in female cancer remains undefined.…”