Detection of joining points for genomic DNA circularization in cell culture

We previously reported on extrachromosomal circular DNA extracted from mouse brain. We attempted to reconfirm the formation of circular DNA from this region in a culture system. From a circular DNA-enriched fraction of a mouse embryonic tumor-derived cell line capable of inducing neuronal differentiation, circular DNA in the same region was isolated by nested inverse polymerase chain reaction as performed previously. We attempted to amplify and identify some junctions that were evidence of circularization. In this analysis, we captured several junctions that indicated circularization in cultured cells induced to differentiate into neurons. We observed that some of the sequences shared the same point of attachment, indicating that there exist genomic sequences that are amenable to binding toward circularization. Cells were X-ray-irradiated to determine whether any transformation occurred in DNA circularization. Consequently, circularization junctions occurred after differentiation-induced stimulation and before and after X-ray irradiation. This finding indicated that circularization junctions can be formed from this region without being inhibited by X-ray irradiation and irrespective of cell differentiation stage. Furthermore, the presence of circular DNA was confirmed in which genomic fragments from different chromosomes were replaced. These findings suggest that extrachromosomal circular DNA contributes to the interchromosomal translocation of genomic fragments.