Table 1 Summary of CRISPR-based DNA methylation editing systems for mammalian cells
From: DNA methylation in melanoma immunotherapy: mechanisms and therapeutic opportunities
System | Effect | Mechanism |
|---|---|---|
dCas9-DNMT3A | gene repression | DNMT3A facilitates de novo DNA methylation, repressing the target gene [86, 91, 99] |
dCas9-DNMT3A3L | gene repression | DNMT3L boosts DNMT3A methylation activity despite itself lacking a catalytic domain, increasing system efficacy [102] |
dCas9-DNMT3A3L-KRAB | gene repression | The KRAB domain is added to the dCas9 system to enhance repression beyond DNMT3A and DNMT3L alone [123] |
dCas9-MQ1 | gene repression | MQ1, an engineered prokaryotic CpG DNA methyltransferase, is fused to dCas9, enhancing methylation activity [124] |
dCas9-sMTase | gene repression | A “split methyltransferase” derived from M.SssI is used, where the two components bind at the target CpG site [96] |
dCas9-scFv-DNMT3A (SunTag-DNMT3A) | gene repression | The SunTag system is used with scFv-DNMT3A (instead of VP64) to recruit multiple DNMT3A proteins to the target region [106] |
dCas9-scFv-DNMT3AL (SunTag-DNMT3AL) | gene repression | The SunTag system is used with scFv-DNMT3AL (instead of VP64) to recruit multiple DNMT3AL proteins to the target region [125] |
dCas9-SunTag system | gene activation | The SunTag protein is linked to dCas9, with VP64-fused scFv added to recruit multiple VP64 copies, enhancing gene activation beyond dCas9-VP64 alone [126, 127] |
dCas9-TET1 | gene activation | TET1 removes methyl groups to activate transcription. The dCas9-TET1 system enables targeted DNA demethylation and gene activation [80, 127] |
dCas9, Tet1-MS2 | gene activation | This system uses Effector-MS2 to recruit multiple TET1 proteins, increasing system efficacy [98] |
dCas9-scFv-TET1 (SunTag-TET1) | gene activation | This system uses SunTag with scFv-TET1 fusion (instead of VP64) to recruit more TET1 proteins, enhancing DNA demethylation [97] |
Casilio-ME | gene activation | The Casilio platform is used to co-deliver TET1 and BER-associated proteins GADD45A or NEIL2 for a higher efficacy than SunTag systems [128] |
dCas9-R2 | gene activation | The dCas9-R2 module specifically binds and inhibits the action of endogenous DNMT1 to prevent local DNA methylation at the target site [129] |
Dual Cas9, Template | gene repression or activation | Excised a 1120 bp CGI from the HPRT1 promoter using dual gRNA-guided Cas9 and replaced it with fully methylated or unmethylated fragments via NHEJ [130] |
dCas9-ROS1 | gene activation | ROS1, a plant-specific DNA glycosylase, directly excises 5mC [131] |
CRISPRoff | gene repression | A novel dCas9-DNMT3A-3LZNF10 KRAB system leads to heritable, persistent gene silencing [88] |
CRISPRon | gene activation | dCas9-TET1 and modified sgRNA with MS2 stem-loop sequences interact with MS2 coat protein fused to the VPR system, to activate silenced genes [88] |