Abstract
Chromatin proteins have expanded the mammalian synthetic biology toolbox by enabling control of active and silenced states at endogenous genes. Others have reported synthetic proteins that bind DNA and regulate genes by altering chromatin marks, such as histone modifications. Previously, we reported the first synthetic transcriptional activator, the “Polycomb-based transcription factor” (PcTF) that reads histone modifications through a protein–protein interaction between the polycomb chromodomain motif and trimethylated lysine 27 of histone H3 (H3K27me3). Here, we describe the genome-wide behavior of the polycomb-based transcription factor fusion protein. Transcriptome and chromatin profiling revealed several polycomb-based transcription factor-sensitive promoter regions marked by distal H3K27me3 and proximal fusion protein binding. These results illuminate a mechanism in which polycomb-based transcription factor interactions bridge epigenomic marks with the transcription initiation complex at target genes. In three cancer-derived human cell lines tested here, some target genes encode developmental regulators and tumor suppressors. Thus, the polycomb-based transcription factor represents a powerful new fusion protein-based method for cancer research and treatment where silencing marks are translated into direct gene activation.
Highlights
Proteins from the gene regulatory complex known as chromatin mediate stable, epigenetic expression states that persist over multiple cell divisions in metazoan tissues
We used an intercalating dye (SYBR green) for Quantitative reverse transcription PCR (qRT-PCR) and observed that CDKN2A was strongly upregulated by Polycomb-based transcription factor” (PcTF) compared to ΔTF in U-2 OS cells.[11]
By including several more known polycomb targets, we have identified other genes that are upregulated by PcTF in all three cell types: IRF8, CADM1, and RUNX3
Summary
Proteins from the gene regulatory complex known as chromatin mediate stable, epigenetic expression states that persist over multiple cell divisions in metazoan tissues. Harnessing the potent gene-regulating functions of chromatin proteins has become a high priority for cancer therapy and tissue engineering. The “histone code” model of chromatin function[1] has strongly influenced work in epigenetic engineering and drug development.[2,3] According to this model, biochemical marks are written onto DNA-bound histone proteins and these marks are read when effector proteins physically interact with the modified histones. Constructive approaches, where synthetic systems are built from chromatin components, are gaining recognition as an important and powerful research method[7] as well as a powerful application for biomedical engineering
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