Fig. 7

Representative working models of targeted demethylation. a With the gDNA, a DNA modification domain, such as TET1, fused to dCas9, led to the erasure of specific DNA methylation.³³³ b GCN4 repeats fused to dCas9, recruited many copies of an anti-GCN4 antibody (scFV)-fused TET1, to amplify demethylation efficiently.³²² c Multiple effectors were used to increase the efficacy of demethylation. The modified gRNA with PUF binding sites, recruited protein fusions of PUF, TET1, and NEIL2 to particular DNA methylation sites. Among these, PUF were used for binding to the modified gRNA, and TET1 oxidated 5mC and NEIL2 worked as a DNA glycosylase to promote DNA demethylation.³²⁴ d Without tethering an effector such as TET1, only gRNA-dCas9 led to specific DNA demethylation, by sterically blocking DNA methyltransferase³³¹