Systematic Comparison of CRISPR-Based Transcriptional Activators Uncovers Gene-Regulatory Features of Enhancer-Promoter Interactions

Nuclease-inactivated CRISPR/Cas-based (dCas-based) systems have emerged as powerful technologies to synthetically reshape the human epigenome and tune gene expression. Despite the straightforward programmability and rapidly increasing adoption of these platforms, the relative potencies, and mechanistic differences between CRISPR/Cas-based transcriptional activators are incompletely characterized, particularly at human enhancer-promoter pairs. Here we systematically compared the transcriptional effects of the most potent and widely used dCas9-based transcriptional activators, as well as a novel activator consisting of dCas9 fused to the catalytic core of the human CBP protein, at human enhancer-promoter pairs. We find that although each system can generally transactivate targeted human loci, the platforms display variable expression levels in different human cell types and further, that the relative efficacies for transcriptional activation vary substantially based upon the effector domain used, recruitment architecture, targeted locus, and the cell type assayed. We also show that when targeted to an enhancer, each dCas9-based activator can induce the production of enhancer RNAs (eRNAs), and that this induction of eRNAs is tightly correlated with downstream mRNA expression from a cognate promoter. Our studies also reveal that different dCas9-based transcriptional activators can deposit histone acetylation at a targeted enhancer, which results in a highly correlated increase in histone acetylation at a downstream promoter. Further, we use dCas9-based transcriptional activators to show that an intrinsic transcriptional and epigenetic reciprocity exists between a testbed human enhancer-promoter pair. Finally, our findings demonstrate that contact frequencies between an enhancer-promoter can highly correlate to transcriptional outputs, which can be manipulated using targeted dCas9-based transcriptional activators also suggest that. Collectively, our study provides new insights into the enhancer-mediated control of gene expression and the use of dCas9-based transcriptional activators at human regulatory elements.