title: Targeted epigenome editing of an endogenous locus with chromatin modifiers is not stably maintained.
abstract: BACKGROUND
  DNA methylation and histone 3 lysine 9 (H3K9) methylation are considered as epigenetic marks that can be inherited through cell divisions. To explore the functional consequences and stability of these modifications, we employed targeted installment of DNA methylation and H3K9 methylation in the vascular endothelial growth factor A (VEGF-A) promoter using catalytic domains of DNA or H3K9 methyltransferases that are fused to a zinc finger protein which binds a site in the VEGF-A promoter.
  
  
  RESULTS
  Expression of the targeted DNA and H3K9 methyltransferases caused dense deposition of DNA methylation or H3K9 di- and trimethylation in the promoter of VEGF-A and downregulation of VEGF-A gene expression. We did not observe positive feedback between DNA methylation and H3K9 methylation. Upon loss of the targeted methyltransferases from the cells, the epigenetic marks, chromatin environment, and gene expression levels returned to their original state, indicating that both methylation marks were not stably propagated after their installment.
  
  
  CONCLUSIONS
  The clear anti-correlation between DNA or H3K9 methylation and gene expression suggests a direct role of these marks in transcriptional control. The lack of maintenance of the transiently induced silenced chromatin state suggests that the stability of epigenetic signaling is based on an epigenetic network consisting of several molecular marks. Therefore, for stable reprogramming, either multivalent deposition of functionally related epigenetic marks or longer-lasting trigger stimuli might be necessary.
authors.affiliation: Institute of Biochemistry, Faculty of Chemistry, Stuttgart University, Pfaffenwaldring 55, 70569 Stuttgart, Germany.
authors.forename: Goran
authors.initials: G
authors.lastname: Kungulovski
authors.affiliation: Institute of Biochemistry, Faculty of Chemistry, Stuttgart University, Pfaffenwaldring 55, 70569 Stuttgart, Germany.
authors.forename: Suneetha
authors.initials: S
authors.lastname: Nunna
authors.affiliation: Dr. Margarete Fischer-Bosch Institute of Clinical Pharmacology, Auerbachstraße 112, 70376 Stuttgart, Germany.|University of Tübingen, Geschwister-Scholl-Platz, 72074 Tübingen, Germany.
authors.forename: Maria
authors.initials: M
authors.lastname: Thomas
authors.affiliation: Dr. Margarete Fischer-Bosch Institute of Clinical Pharmacology, Auerbachstraße 112, 70376 Stuttgart, Germany.|University of Tübingen, Geschwister-Scholl-Platz, 72074 Tübingen, Germany.
authors.forename: Ulrich M
authors.initials: UM
authors.lastname: Zanger
authors.affiliation: Max-Planck-Genomzentrum Köln, Carl-von-Linné-Weg 10, 50829 Köln, Germany.
authors.forename: Richard
authors.initials: R
authors.lastname: Reinhardt
authors.affiliation: Institute of Biochemistry, Faculty of Chemistry, Stuttgart University, Pfaffenwaldring 55, 70569 Stuttgart, Germany.
authors.forename: Albert
authors.initials: A
authors.lastname: Jeltsch