Colson, T. | Search bioRxiv

bioRxiv2025

Dot1L-mediated H3K79 methylation licenses TET-dependent DNA demethylation to establish Treg identity

Foxp3+ regulatory T (Treg) cells rely on DNA demethylation to establish and maintain the gene expression program that defines their identity and suppressive function. Although sustained transcription of Treg-specific genes is known to drive this demethylation, the precise mechanism has remained unclear. Here, we show that Dot1L-catalyzed methylation of histone H3 at lysine 79 (H3K79me) is essential for Treg-specific DNA demethylation in both thymic and induced Treg lineages. H3K79me promotes chromatin activation and recruits TET family DNA demethylases to key regulatory loci. Treg-restricted deletion of Dot1L disrupts locus-specific DNA demethylation, diminishes expression of core Treg genes, and precipitates a fatal, early-onset autoimmune syndrome. Conversely, pharmacologic enhancement of TET activity during differentiation rescues DNA demethylation, restores Treg-gene expression, and reinstates suppressive function even in the presence of Dot1L inhibition. Together, these findings identify a critical epigenetic axis--Dot1L-mediated H3K79 methylation driving TET-dependent DNA demethylation--that safeguards Treg cellular identity, function, and immune homeostasis.

immunology↗

bioRxiv2023

Exploring thienothiadiazine dioxides as isosteric analogues of benzo- and pyridothiadiazine dioxides in the search of new AMPA and kainate receptor positive allosteric modulators

The synthesis and biological evaluation on AMPA and kainate receptors of new examples of 3,4-dihydro-2H-1,2,4-thieno[3,2-e]-1,2,4-thiadiazine 1,1-dioxides is described. The introduction of a cyclopropyl chain instead of an ethyl chain at the 4-position of the thiadiazine ring was found to dramatically improve the potentiator activity on AMPA receptors, with compound 32 (BPAM395) expressing in vitro activity on AMPARs (EC2x = 0.24 {micro}M) close to that of the reference 4-cyclopropyl-substituted benzothiadiazine dioxide 10 (BPAM344). Interestingly, the 4-allyl-substituted thienothiadiazine dioxide 27 (BPAM307) emerged as the most promising compound on kainate receptors being a more effective potentiator than the 4-cyclopropyl-substituted thienothiadiazine dioxide 32 and supporting the view that the 4-allyl substitution of the thiadiazine ring could be more favorable than the 4-cyclopropyl substitution to induce marked activity on kainate receptors versus AMPA receptors. The thieno-analogue 36 (BPAM279) of the clinically tested S18986 (11) was selected for in vivo evaluation in mice as a cognitive enhancer due to a safer profile than 32 after massive per os drug administration. Compound 36 was found to increase the cognition performance in mice at low doses (1 mg/kg) per os suggesting that the compound was well absorbed after oral administration and able to reach the central nervous system. Finally, compound 32 was selected for co-crystallization with the GluA2-LBD (L504Y,N775S) and glutamate to examine the binding mode of thienothiadiazine dioxides within the allosteric binding site of the AMPA receptor. At the allosteric site, this compound established similar interactions as the previously reported BTD-type AMPA receptor modulators. HighlightsO_LIThe study explored AMPA/kainate receptor PAMs belonging to thienothiadiazine dioxides C_LIO_LIThe 4-cyclopropyl-substituted compound 32 was the most potent AMPA receptor modulator C_LIO_LI4-Allyl substitution improved activity and selectivity for kainate receptors C_LIO_LIThe tricyclic compound 36 expressed cognitive improvement in vivo in mice C_LIO_LICompound 32 was co-crystallized with GluA2-LBD to examine receptor binding mode C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=68 SRC="FIGDIR/small/565294v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@fab46corg.highwire.dtl.DTLVardef@15b42dborg.highwire.dtl.DTLVardef@82740eorg.highwire.dtl.DTLVardef@916124_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗