Reactivation of hprt on the inactive X chromosome with DNA demethylating agents.
The C86 line of female embryonal carcinoma cells contains one active and one inactive X chromosome. Following methylnitrosourea mutagenesis, a clone called C86AGM2 was isolated that carries a mutated hprt gene on the active X chromosome. This hprtm allele encodes an HPRT enzyme that has less than 1% normal enzyme activity, is thermolabile, and has an altered isoelectric point. Following treatment with drugs that demethylate DNA, the hprt+ gene from the inactive X chromosome in C86AGM2 cells became active as determined by the appearance of HPRT activity with the thermodenaturation and electrofocusing characteristics of the normal enzyme. No expression of this hprt+ gene occurred if C86AGM2 cells were induced to differentiate prior to DNA demethylation. Stable lines of C86AGM2 cells expressing both the hprtm and hprt+ genes did not inactivate either gene following differentiation.
Hockey
A J
AJ
Department of Medicine, University of Ottawa, Ontario, Canada.
Adra
C N
CN
McBurney
M W
MW
D000818:Animals; D001374:Azacitidine / Q000494:pharmacology; D002454:Cell Differentiation; D004261:DNA Replication; D004303:Dosage Compensation, Genetic; D005786:Gene Expression Regulation; D005796:Genes; D006358:Hot Temperature; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics*; D007525:Isoelectric Focusing; D008745:Methylation; D051379:Mice; D011489:Protein Denaturation; D012333:RNA, Messenger / Q000235:genetics; D014407:Tumor Cells, Cultured; D014960:X Chromosome / Q000502:physiology*
D012333:RNA, Messenger; D007041:Hypoxanthine Phosphoribosyltransferase; D001374:Azacitidine