5-methylcytosine is present in the 5' flanking region of Ha-ras in mouse liver and increases with ageing.
Modifications to DNA-5-methylcytosine (5MeC) content (i.e., alterations in the level of 5MeC) constitute epigenetic events. In general, hypomethylation of a gene is necessary but not sufficient for expression, while methylated genes typically are quiescent. Ha-ras is an oncogene commonly implicated in murine liver tumorigenesis, often, though not always, involving mutation. A PCR-based approach using pre-PCR digestion with methylation-sensitive enzymes was employed to determine the 5MeC content of the 5' flanking region of this gene in (i) B6C3F1 and C57BL/6 mouse liver from young animals (4 months old) and (ii) B6C3F1 mouse liver from aged animals (24 months old). Two segments of the 5' flanking region of Ha-ras were examined. We demonstrate the presence of 5MeC in a portion of the 5' flanking region of Ha-ras that does not share characteristics of a CpG island, while a region that shares CpG island characteristics is primarily unmethylated. Differences in methylation status in these areas of Ha-ras were not observed between B6C3F1 and C57BL/6 mouse livers. Increases in methylation status were observed with ageing in B6C3F1 mouse liver. These data provide a role for methylation in regulating Ha-ras expression in mouse liver. Ha-ras in human liver has been reported to be unmethylated. There are substantial sequence differences in a key region of the 5' flanking region of Ha-ras in mice as compared to humans. These differences in DNA methylation and sequence may, in part, provide a basis for the frequent involvement of Ha-ras in mouse liver tumors and its virtual lack of involvement in human tumors.
Counts
J L
JL
Department of Pharmacology and Toxicology, Michigan State University, East Lansing 48824, USA.
Kaznowski
J M
JM
McClain
R M
RM
Goodman
J I
JI
D044503:5-Methylcytosine; D000375:Aging / Q000235:genetics*; D000818:Animals; D003596:Cytosine / Q000031:analogs & derivatives* / Q000032:analysis / Q000378:metabolism; D019175:DNA Methylation; D004262:DNA Restriction Enzymes / Q000378:metabolism; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D011905:Genes, ras; D006801:Humans; D008099:Liver / Q000737:chemistry* / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D016133:Polymerase Chain Reaction
D044503:5-Methylcytosine; D003596:Cytosine; D004262:DNA Restriction Enzymes; C031851:endodeoxyribonuclease HaeII; D015252:Deoxyribonucleases, Type II Site-Specific