It has been known for a long time that the cytosine base can be modified to produce a new nucleotide, identified as 5-methylcytosine (mC). In normal cells, mC is correctly distributed into the genome, but in many diseases including life-threatening cancers, its pattern is profoundly perturbed. In 2009, Anjana Rao, published that certain proteins, known as the TET enzymes, are capable of removing mC by further oxidizing it to 5-hydroxymethylcytosine (hmC). This original article, cited more than 1200 times, has led to a great expansion in our understanding of DNA methylation. Such recent publications expanded this knowledge by showing that the TETs successively oxidize hmC to 5-formylcytosine (fC) and 5-carboxylcytosine (caC). <p>These oxidized methylcytosines have been implicated in several mechanisms of DNA demethylation, including “active” demethylation through base excision repair, and “passive” demethylation via successive rounds of DNA replication. In addition, DNA hydroxymethylation is thought to be involved in a wide range of diseases, and a marked decrease of hmC seems to be a “hallmark” of many cancers. <p>However, little is known about the regulation of their modes of action. It is tempting to speculate that these proteins interact with a plethora of factors to elicit coordinated biological functions. Likewise, they might be regulated by environment, which in certain situations, could alter the hydroxymethylome landscape, and lead to cellular malfunction and diseases.<p>In the first study, we pursued a large, unbiased screen of the TET interactome, and discovered that TET2 and TET3 interact with the O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT). OGT is a glycosyltransferase that adds N-acetylglucose moieties on various proteins, including histone H2B, expanding therefore the “histone code”. We further discovered that the TET-OGT association seems to enhance OGT activity and to potentiate glycosylation and stabilization of SET1/COMPASS, a complex that is responsible for the global deposition of the H3K4me3 histone mark that “decorates” active promoters. Finally, we could confirm a decreased genome-wide H3K4me3 deposition in a model of acute myeloid leukemia mutated for TET2, suggesting that the TET-OGT link is implicated in Health and Disease.<p>In the second study, we looked at the impact of the environment on TET activity and on cellular hydroxymethylomes. We focused on oxidative stress assaults that are known to be involved in inflammation, a mediator of cancer and neurodegenerative diseases. We observed a significant decrease of hmC in cell lines treated with various oxidant stressors, likely due to a direct inactivation of the TETs catalytic domain. Moreover, gene ontology analysis of differentially hydroxymethylated regions (dhMRs), profiled by deep-sequencing on treated vs non-treated cells, highlighted pathways involved in oxidative stress response. The implication of TETs in oxidative stress response was further emphasized by a decreased proliferation of TET1-depleted cells when they are treated with oxidant stressors. Importantly, those results were confirmed in mice knockout for the major antioxidant enzymes GPx1 and GPx2. <p>In conclusion, the work of this thesis contributed to better understand the modes of action of the TET proteins, through (1) direct interaction with OGT, and (2) via direct regulation by oxidative-stress-associated molecules, and we hope that these results will bring new insights to better understand these fascinating enzymes. <p> / Doctorat en Sciences biomédicales et pharmaceutiques / info:eu-repo/semantics/nonPublished
| Identifer | oai:union.ndltd.org:ulb.ac.be/oai:dipot.ulb.ac.be:2013/209201 |
| Date | 01 October 2014 |
| Creators | Delatte, Benjamin |
| Contributors | Fuks, François, Vassart, Gilbert, Abramowicz, Marc, Christophe, Daniel, Goriely, Stanislas |
| Publisher | Universite Libre de Bruxelles, Université libre de Bruxelles, Faculté de Médecine – Sciences biomédicales, Bruxelles |
| Source Sets | Université libre de Bruxelles |
| Language | English |
| Detected Language | English |
| Type | info:eu-repo/semantics/doctoralThesis, info:ulb-repo/semantics/doctoralThesis, info:ulb-repo/semantics/openurl/vlink-dissertation |
| Format | No full-text files |
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