Fd. Araujo et al., Identification of initiation sites for DNA replication in the human dnmt1 (DNA-methyltransferase) locus, J BIOL CHEM, 274(14), 1999, pp. 9335-9341
Vertebrates have developed multiple mechanisms to coordinate the replicatio
n of epigenetic and genetic information. Dnmt1 encodes the maintenance enzy
me DNA-methyltransferase, which is responsible for propagating the DNA meth
ylation pattern and the epigenetic information that it encodes during repli
cation. Direct sequence analysis and bisulfite mapping of the 5' region of
DNA-methyltransferase 1 (dnmt1) have indicated the presence of many sequenc
e elements associated with previously characterized origins of DNA replicat
ion. This study tests the hypothesis that the dnmt1 region containing these
elements is an origin of replication in human cells. First, we demonstrate
that a vector containing this dnmt1 sequence is able to support autonomous
replication when transfected into HeLa cells. Second, using a gel retardat
ion assay, we show that it contains a site for binding of origin-rich seque
nces binding activity, a recently purified replication protein. Finally, us
ing competitive polymerase chain reaction, we show that replication initiat
es in this region in vivo, Based on these lines of evidence, we propose tha
t initiation sites for DNA replication are located between the first intron
and exon 7 of the human dnmt1 locus.