DNA methylation of the extraembryonic tissues: an in situ study on human metaphase chromosomes

DNA methylation level and pattern of human metaphase chromosomes from extraembryonic tissues (chorionic villi and placental fibroblasts) were analysed in situ. The DNA methylation global level of these tissues was studied by comparing them with the one observed in fetal fibroblasts and adult lymphocytes. In order to assess the tissue specificity and significance of the observed differences, chromosomal preparations were then treated in parallel. They were first stained with distamycin A/DAPI and pictured, then treated with immunofluorescent staining using monoclonal antibodies raised against 5-methylcytosine. Compared with metaphases from lymphocytes or placental and fetal fibroblasts, distamycin-A/DAPI stained metaphases and constitutive heterochromatic regions with very similar intensities. In contrast, in chorionic villi, the immunofluorescent intensities revealing the presence of 5-methylcytosine was much duller than in the other tissues. In addition, in both chorionic villi and placental fibroblasts, large differences were observed between various chromosome structures within individual metaphases. In particular, the secondary constriction of chromosome 9, the distal segment of chromosome Y and the short arms of acrocentric chromosomes exhibited a much lower staining than the one observed for the secondary constrictions of chromosome 1 and 16 of the same metaphases. Because all these structures are known to be deeply methylated in other somatic tissues, this suggests that in extraembryonic tissues DNA methylation level remained hypomethylated and the pattern is under precise control.