Abstract

Fusion proteins that specifically interact with biochemical marks on chromosomes represent a new class of synthetic transcriptional regulators that decode cell state information rather than DNA sequences. In multicellular organisms, information relevant to cell state, tissue identity, and oncogenesis is often encoded as biochemical modifications of histones, which are bound to DNA in eukaryotic nuclei and regulate gene expression states. We have previously reported the development and validation of the "polycomb-based transcription factor" (PcTF), a fusion protein that recognizes histone modifications through a protein-protein interaction between its polycomb chromodomain (PCD) motif and trimethylated lysine 27 of histone H3 (H3K27me3) at genomic sites. We demonstrated that PcTF activates genes at methyl-histone-enriched loci in cancer-derived cell lines. However, PcTF induces modest activation of a methyl-histone associated reporter compared to a DNA-binding activator. Therefore, we modified PcTF to enhance its binding avidity. Here, we demonstrate the activity of a modified regulator called Pc2TF, which has two tandem copies of the H3K27me3-binding PCD at the N-terminus. Pc2TF has a smaller apparent dissociation constant value in vitro and shows enhanced gene activation in HEK293 cells compared to PcTF. These results provide compelling evidence that the intrinsic histone-binding activity of the PCD motif can be used to tune the activity of synthetic histone-binding transcriptional regulators.

Original languageEnglish (US)
Pages (from-to)842-852
Number of pages11
JournalACS Synthetic Biology
Volume7
Issue number3
DOIs
StatePublished - Mar 16 2018

Fingerprint

Transcription factors
Histones
Chromatin
Transcription Factors
Histone Code
Proteins
DNA
Fusion reactions
Genes
Chemical activation
HEK293 Cells
DNA sequences
Chromosomes
Gene expression
Transcriptional Activation
Lysine
Carcinogenesis
Cells
Tissue
Gene Expression

Keywords

  • chromatin
  • epigenetics
  • histone
  • protein engineering
  • synthetic biology
  • transcription factor

ASJC Scopus subject areas

  • Biomedical Engineering
  • Biochemistry, Genetics and Molecular Biology (miscellaneous)

Cite this

Tandem Histone-Binding Domains Enhance the Activity of a Synthetic Chromatin Effector. / Tekel, Stefan J.; Vargas, Daniel A.; Song, Lusheng; LaBaer, Joshua; Caplan, Michael; Haynes, Karmella.

In: ACS Synthetic Biology, Vol. 7, No. 3, 16.03.2018, p. 842-852.

Research output: Contribution to journalArticle

Tekel, Stefan J. ; Vargas, Daniel A. ; Song, Lusheng ; LaBaer, Joshua ; Caplan, Michael ; Haynes, Karmella. / Tandem Histone-Binding Domains Enhance the Activity of a Synthetic Chromatin Effector. In: ACS Synthetic Biology. 2018 ; Vol. 7, No. 3. pp. 842-852.
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