LINE-1 methylation status of endogenous DNA double-strand breaks.
DNA methylation and the repair of DNA double-strand breaks (DSBs) are important processes for maintaining genomic integrity. Although DSBs can be produced by numerous agents, they also occur spontaneously as endogenous DSBs (EDSBs). In this study, we evaluated the methylation status of EDSBs to determine if there is a connection between DNA methylation and EDSBs. We utilized interspersed repetitive sequence polymerase chain reaction (PCR), ligation-mediated PCR and combined bisulfite restriction analysis to examine the extent of EDSBs and methylation at long interspersed nuclear element-1 (LINE-1) sequences nearby EDSBs. We tested normal white blood cells and several cell lines derived from epithelial cancers and leukemias. Significant levels of EDSBs were detectable in all cell types. EDSBs were also found in both replicating and non-replicating cells. We found that EDSBs contain higher levels of methylation than the cellular genome. This hypermethylation is replication independent and the methylation was present in the genome at the location prior to the DNA DSB. The differences in methylation levels between EDSBs and the rest of the genome suggests that EDSBs are differentially processed, by production, end-modification, or repair, depending on the DNA methylation status.
Pornthanakasem
Wichai
W
Center of Excellence in Molecular Genetics of Cancer and Human Diseases, Department of Anatomy, Faculty of Medicine, Chulalongkorn University, Bangkok 10330, Thailand.
Kongruttanachok
Narisorn
N
Phuangphairoj
Chutipa
C
Suyarnsestakorn
Chotika
C
Sanghangthum
Taweap
T
Oonsiri
Sornjarod
S
Ponyeam
Wanpen
W
Thanasupawat
Thatchawan
T
Matangkasombut
Oranart
O
Mutirangura
Apiwat
A
D045744:Cell Line, Tumor; D053903:DNA Breaks, Double-Stranded; D019175:DNA Methylation; D004261:DNA Replication; D015894:Genome, Human; D006801:Humans; D020084:Long Interspersed Nucleotide Elements; D016133:Polymerase Chain Reaction / Q000379:methods