Gene silencing in mammalian cells by uptake of 5-methyl deoxycytidine-5'-triphosphate.
Chinese hamster ovary (CHO) cells were subjected to electroporation in the presence of 5-methyl deoxycytidine-triphosphate. This treatment increases by 10 to 100-fold the frequency of cells lacking thymidine kinase, hypoxanthine-guanine phosphoribosyltransferase, or adenine phosphoribosyltransferase. The inactivation of the genes coding for these enzymes is thought to occur following the direct incorporation of the methylated nucleotide triphosphate into DNA. The enzyme-deficient clones were stable, but almost all were reactivated at high frequency by the demethylating agent 5-azacytidine, to produce derivatives with enzyme activity. The results indicate that there is a direct relationship between DNA methylation and gene silencing.
Holliday
R
R
CSIRO Division of Biomolecular Engineering, Laboratory for Molecular Biology, North Ryde, NSW, Australia.
Ho
T
T
D000228:Adenine Phosphoribosyltransferase / Q000235:genetics; D000818:Animals; D001374:Azacitidine / Q000494:pharmacology; D001973:Bromodeoxyuridine / Q000494:pharmacology; D016466:CHO Cells; D006224:Cricetinae; D004247:DNA / Q000378:metabolism*; D003845:Deoxycytosine Nucleotides / Q000494:pharmacology*; D004351:Drug Resistance; D005786:Gene Expression Regulation / Q000187:drug effects; D005819:Genetic Markers; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics; D013937:Thymidine Kinase / Q000235:genetics; D014960:X Chromosome
D003845:Deoxycytosine Nucleotides; D005819:Genetic Markers; C034817:5-methyldeoxycytidine triphosphate; D004247:DNA; D000228:Adenine Phosphoribosyltransferase; D007041:Hypoxanthine Phosphoribosyltransferase; D013937:Thymidine Kinase; D001973:Bromodeoxyuridine; D001374:Azacitidine