KRAB-Induced Heterochromatin Effectively Silences PLOD2 Gene Expression in Somatic Cells and is Resilient to TGFβ1 Activation

Rutger A F Gjaltema, Désirée Goubert, Christian Huisman, Consuelo Del Pilar García Tobilla, Mihály Koncz, Pytrick G Jellema, Dandan Wu, Uilke Brouwer, Antal Kiss, Pernette J Verschure, Ruud A Bank, Marianne G Rots*

*Corresponding author for this work

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Abstract

Epigenetic editing, an emerging technique used for the modulation of gene expression in mammalian cells, is a promising strategy to correct disease-related gene expression. Although epigenetic reprogramming results in sustained transcriptional modulation in several in vivo models, further studies are needed to develop this approach into a straightforward technology for effective and specific interventions. Important goals of current research efforts are understanding the context-dependency of successful epigenetic editing and finding the most effective epigenetic effector(s) for specific tasks. Here we tested whether the fibrosis- and cancer-associated PLOD2 gene can be repressed by the DNA methyltransferase M.SssI, or by the non-catalytic Krüppel associated box (KRAB) repressor directed to the PLOD2 promoter via zinc finger- or CRISPR-dCas9-mediated targeting. M.SssI fusions induced de novo DNA methylation, changed histone modifications in a context-dependent manner, and led to 50%-70% reduction in PLOD2 expression in fibrotic fibroblasts and in MDA-MB-231 cancer cells. Targeting KRAB to PLOD2 resulted in the deposition of repressive histone modifications without DNA methylation and in almost complete PLOD2 silencing. Interestingly, both long-term TGFβ1-induced, as well as unstimulated PLOD2 expression, was completely repressed by KRAB, while M.SssI only prevented the TGFβ1-induced PLOD2 expression. Targeting transiently expressed dCas9-KRAB resulted in sustained PLOD2 repression in HEK293T and MCF-7 cells. Together, these findings point to KRAB outperforming DNA methylation as a small potent targeting epigenetic effector for silencing TGFβ1-induced and uninduced PLOD2 expression.

Original languageEnglish
Article number3634
Number of pages24
JournalInternational Journal of Molecular Sciences
Volume21
Issue number10
DOIs
Publication statusPublished - 21-May-2020

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