PT - JOURNAL ARTICLE AU - Verena Thormann AU - Laura V Glaser AU - Maika C Rothkegel AU - Marina Borschiwer AU - Melissa Bothe AU - Alisa Fuchs AU - Sebastiaan H Meijsing TI - Expanding the repertoire of glucocorticoid receptor target genes by engineering genomic response elements AID - 10.26508/lsa.201800283 DP - 2019 Apr 01 TA - Life Science Alliance PG - e201800283 VI - 2 IP - 2 4099 - https://www.life-science-alliance.org/content/2/2/e201800283.short 4100 - https://www.life-science-alliance.org/content/2/2/e201800283.full SO - Life Sci. Alliance2019 Apr 01; 2 AB - The glucocorticoid receptor (GR), a hormone-activated transcription factor, binds to a myriad of genomic binding sites yet seems to regulate a much smaller number of genes. Genome-wide analysis of GR binding and gene regulation has shown that the likelihood of GR-dependent regulation increases with decreased distance of its binding to the transcriptional start site of a gene. To test if we can adopt this knowledge to expand the repertoire of GR target genes, we used CRISPR/Cas-mediated homology-directed repair to add a single GR-binding site directly upstream of the transcriptional start site of each of four genes. To our surprise, we found that the addition of a single GR-binding site can be enough to convert a gene into a GR target. The gain of GR-dependent regulation was observed for two of four genes analyzed and coincided with acquired GR binding at the introduced binding site. However, the gene-specific gain of GR-dependent regulation could not be explained by obvious differences in chromatin accessibility between converted genes and their non-converted counterparts. Furthermore, by introducing GR-binding sequences with different nucleotide compositions, we show that activation can be facilitated by distinct sequences without obvious differences in activity between the GR-binding sequence variants we tested. The approach to use genome engineering to build genomic response elements facilitates the generation of cell lines with tailored repertoires of GR-responsive genes and a framework to test and refine our understanding of the cis-regulatory logic of gene regulation by testing if engineered response elements behave as predicted.