High-fidelity CRISPR/Cas9-based gene-specific hydroxymethylation rescues gene expression and attenuates renal fibrosisXu, X., Tan, X., Tampe, B., Wilhelmi, T., Hulshoff, M. S., Saito, S., Moser, T., Kalluri, R., Hasenfuss, G., Zeisberg, E. M. & Zeisberg, M., 29-Aug-2018, In : Nature Communications. 9, 15 p., 3509.
Research output: Contribution to journal › Article › Academic › peer-review
While suppression of specific genes through aberrant promoter methylation contributes to different diseases including organ fibrosis, gene-specific reactivation technology is not yet available for therapy. TET enzymes catalyze hydroxymethylation of methylated DNA, reactivating gene expression. We here report generation of a high-fidelity CRISPR/Cas9-based gene-specific dioxygenase by fusing an endonuclease deactivated high-fidelity Cas9 (dHFCas9) to TET3 catalytic domain (TET3CD), targeted to specific genes by guiding RNAs (sgRNA). We demonstrate use of this technology in four different anti-fibrotic genes in different cell types in vitro, among them RASAL1 and Klotho, both hypermethylated in kidney fibrosis. Furthermore, in vivo lentiviral delivery of the Rasal1-targeted fusion protein to interstitial cells and of the Klotho-targeted fusion protein to tubular epithelial cells each results in specific gene reactivation and attenuation of fibrosis, providing gene-specific demethylating technology in a disease model.
|Number of pages||15|
|Publication status||Published - 29-Aug-2018|
- TARGETED DNA DEMETHYLATION, TUMOR-SUPPRESSOR, PROMOTER METHYLATION, CRISPR-CAS9 SYSTEM, KLOTHO EXPRESSION, GASTRIC-CANCER, BREAST-CANCER, KIDNEY, RASAL1, ACTIVATION