Cell and Molecular Biology of DNA Methyltransferase 1 - International Review of Cell and Molecular Biology - page 31

functions of the deaminases AID and MBD4 (deaminases) and glycosylases
such as TDG (thymine DNA glycosylase).
Another proposed active demethylation mechanism for mammals starts
with the generation and recognition of 5-hmC in DNA ( Zhang et al.,
2012a,b ). 5-hmC is derived from the oxidation of 5-methylcytosine.
5-hmC can be generated in cells by the activity of TET1, a homolog of
JBP1 and JBP2 and related TET proteins TET2 and TET3 ( Tan and Shi,
2012 ). Roles of TET proteins in demethylation has been suggested by
experiments in which genomic methylation is observed in association with
overexpression of one or more of the TET proteins. As is the case with pro-
posed deamination-initiated demethylation mechanisms, DNA glycosylases
and GADD45 proteins may also play roles in the proposed 5-hmC-mediated
demethylation mechanisms ( Niehrs and Sch¨fer, 2012 ). The primary evi-
dence for this is the observation that overexpression of GADD45 proteins
and glycosylases in zebrafish embryos leads to reduction in genomic meth-
ylation. In Xenopus , GADD45a may initiate demethylation along with XPG
(NR protein) ( Barreto et al., 2007 ).
Although there is experimental evidence obtained from in vitro studies as
well as in vivo animal studies supporting the existence of different active CpG
demethylation mechanisms, there is concern as to whether these mecha-
nisms, should they be active in vivo , could alone or collectively account
for the very significant loss of genomic methylation seen at certain develop-
mental times. An active demethylation mechanism based on deamination of
5-methyl-cytosine to thymine generates potentially mutagenic T:G
mismatches that, if not corrected properly to C:G matches, would lead to
T:A matches or point mutations. Because of the proposed genomic scales
at which this mechanism would act, T:G mismatches would have to be very
accurately corrected to ensure that the mechanism leads to demethylation
and not an extraordinarily high frequency of point mutations (cytosine to
thymine transitions). Although the GADD45A protein has been proposed
to be involved in active demethylation, Engel et al. (2009) investigated
Gadd45a

/ mice and found no effects on DNA methylation.

Gadd45b mice show defects in neural progenitor proliferation and

dendritic growth, and there is some loss of methylation, but the possible role
of demethylation was not investigated biochemically ( Ma et al., 2009 ).
Similarly, mouse knockouts of catalysts of the proposed reactions show some
increase in genomic methylation in cells normally undergoing demethyla-
tion, but the changes are minimal, raising doubts about the precise roles
of

/

these entities in the proposed active demethylation processes.