(A and B) Overexpression of AID results in a decrease in the abundance of 5hmCs in genomic DNA samples from HEK 293 cells as monitored by immunoblotting (A) and ELISA (B) using anti-5hmC antibodies.
(C) Effects of AID on demethylation of 5hmC-GFP and 5mC-GFP DNA. Values represent mean ± SEM (n = 3; *: P < 0.05; #: P > 0.1; Student’s t-test)
(D) Effects of overexpression of AID/APOBEC deaminases on 5hmC demethylation as measured by bisulfite sequencing. Values represent mean ± SEM (n = 3; **: P < 0.01; *: P < 0.05; Student’s t-test).
(E) Immunoblotting analysis of 5hmU levels in HEK293 cells after TET1 expression or transfection of 5hmC-GFP DNA, with or without the expression of 5hmU glycosylase SMUG1.
(F) An oxidation-deamination-BER model of TET1-induced active DNA demethylation in mammalian cells.