title: Silencing and activating heterochromatin
creator: Rademacher, Anne
subject: ddc-570
subject: 570 Life sciences
description: Repetitive, gene-poor regions termed heterochromatin are typically transcriptionally silent and mostly located at specific chromosomal loci such as (peri)centromeres and telomeres, but are also found interspersed throughout the rest of the genome. Heterochromatin silencing is associated with epigenetic features such as DNA methylation and posttranslational modification of histone tails, e. g. trimethylation of histone 3 lysine 9 (H3K9me3), which is set by the histone methyltransferases SUV39H1 and SUV39H2 among others. However, the mechanisms underlying establishment and stability of repressive histone marks as well as their contribution to transcriptional silencing remain unclear. Here, I show that the SUV39H histone lysine methyltransferases recognize heterochromatin downstream of the methyl-DNA binding protein MECP2 and remain chromatin-bound throughout the cell cycle, linking these enzymes to the specificity and memory of heterochromatin. In addition, SUV39H isoforms displayed distinct non-redundant functions. SUV39H2 contributed more to establishing H3K9me3 than SUV39H1 in mouse embryonic fibroblasts and loss of SUV39H-dependent H3K9me3 was not sufficient for transcriptional derepression. Since tools to resolve the kinetics of transcriptional activation with high spatial and temporal resolution are currently scarce, I devised the Blue Light-Induced Chromatin Recruitment (BLInCR) tool to dissect the activation of the repressed heterochromatin state. I used BLInCR to rapidly and reversibly target the viral transactivator VP16 to an integrated reporter in the U2OS 2-6-3 cell line and to measure the activation kinetics of an initially repressed reporter gene locus. First transcripts were detectable within minutes after triggering the relocalization of VP16 and activation progressed in two phases. Thus, even a strong activator was not able to activate silenced gene clusters in one step. The BLInCR tool introduced here enables the use of complex activation patterns, which will also be valuable to assess the effect of repeated activation and, more generally, to probe the stability of chromatin states to understand the regulation of epigenetic memory. Taken together, these results indicate that only some genes are susceptible to SUV39H- and H3K9me3-mediated repression and that the heterochromatin state can be overwritten by strong, targeted activators. The insights gained within this study on the mechanisms of silencing and activating heterochromatin can help understand heterochromatic deregulation. The latter is frequently observed in disease, e. g. in cancer, and thus, the heterochromatin network might harbor promising targets for epigenetic drug development.
date: 2018
type: Dissertation
type: info:eu-repo/semantics/doctoralThesis
type: NonPeerReviewed
format: application/pdf
identifier: https://archiv.ub.uni-heidelberg.de/volltextserverhttps://archiv.ub.uni-heidelberg.de/volltextserver/23768/1/RademacherAnne_PhDthesis_2017.pdf
identifier: DOI:10.11588/heidok.00023768
identifier: urn:nbn:de:bsz:16-heidok-237687
identifier:   Rademacher, Anne  (2018) Silencing and activating heterochromatin.  [Dissertation]     
relation: https://archiv.ub.uni-heidelberg.de/volltextserver/23768/
rights: info:eu-repo/semantics/openAccess
rights: http://archiv.ub.uni-heidelberg.de/volltextserver/help/license_urhg.html
language: eng