In vitro correction of G o T mispairs to G o C pairs in nuclear extracts from human cells

Publisher Website
Google Scholar
Abstract
IN differentiated cells, only a specific subset of genes is expressed. Recently, several genes have been shown to be transcriptionally inactivated by methylation of cytosine residues, mainly within their promoter sequences1–4. Spontaneous hydrolytic deamination of 5-methylcytosine to thymine5, which has been estimated to generate up to 12 G-T mismatched base pairs in the human genome per day6,7, could have a deleterious effect on the expression of such genes. We recently reported that mammalian cells possess a specific repair pathway, which counteracts the mutagenic effects of this deamination by correcting G ṁ T mismatches almost exclusively to G ṁ C pairs8,9. We show here that, in nuclear extracts from HeLa cells, this repair is mediated by excision of the aberrant thymidine monophosphate residue, followed by gap-filling to generate a G ṁ C pair. We also provide preliminary evidence that the initial step of this process involves a DNA glycosylase.