Rapid methylation of genomic DNA in proliferating T-cells from bovine thymocytes.
Two subpopulations of bovine calf thymus cells were separated by buoyant density centrifugation. The low-density cells (L-cells) showed high response to T-cell mitogens, while the high density cells (H-cells) did not. The DNA-metabolizing enzyme activities were elevated 10-fold in L-cells in comparison with those in H-cells. L-cells contained a DNA-replicating complex, DNA replitase, while H-cells did not. These observations suggested that L-cells were proliferating, mature T-cells and H-cells were dying intrathymic cells. A DNA methyltransferase was associated with DNA replitase in L-cells. In order to determine whether replitase-associated DNA methyltransferase functions at replicating regions, the methylation pattern of genomic DNA of L-cells was compared with that of H-cells. No significant difference was found in the extent of CpG dinucleotide methylation, and in the location of mC in the satellite I DNA sequence as identified by Southern hybridization and direct sequencing. Thus the majority of methylation patterns of genomic DNA did not change during T-cell development in the thymus. The results indicated that the methylation patterns were rapidly maintained in proliferating T-cells. Although methods employed in the present study might not be sensitive enough to detect transient hemimethylation, it is suggested that the rapid methylation might be catalyzed, albeit not completely, by a DNA methyltransferase associated with the DNA replitase complex.
Sano
H
H
Biotechnology Institute, Akita Prefectural College of Agriculture, Japan.
Noguchi
H
H
Kodama
I
I
Itoh
T
T
D000818:Animals; D002417:Cattle; D002467:Cell Nucleus / Q000201:enzymology; D004247:DNA / Q000235:genetics*; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000378:metabolism; D004261:DNA Replication; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D005796:Genes; D008213:Lymphocyte Activation; D008745:Methylation; D009097:Multienzyme Complexes / Q000378:metabolism; D013601:T-Lymphocytes / Q000145:classification / Q000276:immunology / Q000378:metabolism*; D013937:Thymidine Kinase / Q000378:metabolism; D013950:Thymus Gland / Q000378:metabolism
D009097:Multienzyme Complexes; C037117:replitase; D004247:DNA; D004248:DNA (Cytosine-5-)-Methyltransferases; D013937:Thymidine Kinase; D004259:DNA-Directed DNA Polymerase