2013/08/27 by Albert W Cheng, Albert W. Cheng, Haoyi Wang +11 · 2 citations
Biochemistry, Genetics and Molecular Biology · #Advanced biosensing and bioanalysis techniques #CRISPR and Genetic Engineering #RNA Interference and Gene Delivery
paper · pdf · doi:10.1038/cr.2013.122
openalex publication_date 2013/08/27 · openalex created_date 2025/10/10 · openalex updated_date 2026/07/30
Technologies allowing for specific regulation of endogenous genes are valuable for the study of gene functions and have great potential in therapeutics. We created the CRISPR-on system, a two-component transcriptional activator consisting of a nuclease-dead Cas9 (dCas9) protein fused with a transcriptional activation domain and single guide RNAs (sgRNAs) with complementary sequence to gene promoters. We demonstrate that CRISPR-on can efficiently activate exogenous reporter genes in both human and mouse cells in a tunable manner. In addition, we show that robust reporter gene activation in vivo can be achieved by injecting the system components into mouse zygotes. Furthermore, we show that CRISPR-on can activate the endogenous IL1RN, SOX2, and OCT4 genes. The most efficient gene activation was achieved by clusters of 3-4 sgRNAs binding to the proximal promoters, suggesting their synergistic action in gene induction. Significantly, when sgRNAs targeting multiple genes were simultaneously introduced into cells, robust multiplexed endogenous gene activation was achieved. Genome-wide expression profiling demonstrated high specificity of the system.