Cell and Molecular Biology of DNA Methyltransferase 1 - International Review of Cell and Molecular Biology - page 9

DNMT1. That is, hemimethylated DNA that appears after the first cycle of
DNA replication in the absence of DNMT1 undergoes further cycles of rep-
lication and with each cycle the accumulation of a greater proportion of
unmethylated DNA and a lesser proportion of hemimethylated DNA.
In vivo studies of Dnmt1 gene mutations in mice support these ES-cell
findings—embryos developing in the absence of DNMT1 protein
express de novo methyltransferase DNMT3A and DNMT3B, yet have
greater than an 80% reduction in the normal level of genomic methylation
( Li et al., 1992 ).
The effect of removal of oocyte-derived DNMT1o protein in preim-
plantation embryos on inherited methylation associated with genomic
imprints is particularly insightful in understanding the physiological role
of DNMT1. DNMT1o is a maternal-effect protein that is expressed in
the nucleus only at the 8-cell stage of development, suggesting that it is
active on nuclear genomic DNA at only a single cell cycle ( Mertineit
et al., 1998 ). Removal of this protein leads to, on average, a 50% reduction
in imprinted DNAmethylation in the fetus, a finding that is completely con-
sistent with a maintenance methylation role for DNMT1o in blastomeres of
8-cell embryos ( Cirio et al., 2008b; Howell et al., 2001 ).
3. BASIC MOLECULAR MECHANISMS
OF MAINTENANCE METHYLATION
3.1. Basic chemistry of enzymatic reaction
At its core, the mechanism of maintenance methylation transfers a methyl
group from S -adenosyl methionine to the carbon at the fifth position in
the pyrimidine ring of cytosine. All cytosine methyltransferases, whether
they are primarily de novo or maintenance methyltransferase have this prop-
erty. This basic mechanism as described below is found in both plants
and animals.
The basic chemical mechanism of the transfer of a methyl group from the
cofactor S -adenosyl-methionine (SAM) to a cytosine base is now well
established. This is described in Fig. 1.3 . In brief, the likely mechanism can
be described as four discrete reaction steps ( Santi et al., 1983 ). In step 1, there
is a thiolate attack by the enzyme at position C6 of cytosine. The accompa-
nying protonation of the N3 position is mediated by the highly conserved
ENV peptide motif in the methyltransferase's catalytic domain. Step 2 is an
enamine attack on the methyl group of S -adenosyl methionine. Step 3 is