Differentiation of trophoblast lineage is associated with DNA methylation and demethylation

Our previous study has shown that the placenta and kidney had different genomic methylation patterns regarding CpG island loci detected by restriction landmark genomic scanning (RLGS). To investigate whether differentiation involves changes in DNA methylation, we analyzed the rat Rcho-1 cell line, which retains trophoblast cell features and differentiates from stem cells into trophoblast giant cells in vitro. By RLGS, a total of 1,232 spots were identified in the Rcho-1 stem and differentiated giant cells. Four spots (0.3%) were detected only in giant cells, implying that the loci were originally methylated, but became demethylated during differentiation. Another four spots (0.3%) were detected only in stem cells, implying that these loci, originally unmethylated, became methylated during differentiation. DNAs from three loci that became methylated during differentiation were cloned and sequenced. All showed high homologies with expressed sequence tags (ESTs) or with genomic DNA of other species, suggesting that these loci are biologically important. Thus, the eight differentially methylated loci should be good tools to study epigenetic modification specific to differentiation of trophoblast giant cells.