Formation of carcinogenic chromosomal rearrangements in human thyroid cells after induction of double-strand DNA breaks by restriction endonucleases

Abstract

Ionizing radiation (IR) exposure increases the risk of thyroid cancer and other cancer types. Chromosomal rearrangements, such asRET/PTC, are characteristic features of radiation-associated thyroid cancer and can be induced by radiationin vitro. IR causes double-strand breaks (DSBs), suggesting that such damage leads toRET/PTC, but the rearrangement mechanism has not been established. To study the mechanism, we explored the possibility of inducingRET/PTCby electroporation of restriction endonucleases (REs) into HTori-3 human thyroid cells. We used five REs, which induced DSB in a dose-dependent manner similar to that seen with IR. Although all but one RE caused DSB in one or more of the three genes involved inRET/PTC, rearrangement was detected only in cells electroporated with either PvuII (25 and 100 U) or StuI (100 and 250 U). The predominant rearrangement type wasRET/PTC3, which is characteristic of human thyroid cancer arising early after Chernobyl-related radioactive iodine exposure. Both enzymes that producedRET/PTChad restriction sites only in one of the two fusion partner genes. Moreover, the two enzymes that producedRET/PTChad restriction sites present in clusters, which was not the case for RE that failed to induceRET/PTC. In summary, we establish a model of DSB induction by RE and report for the first time the formation of carcinogenic chromosomal rearrangements, predominantlyRET/PTC3, as a result of DSB produced by RE. Our data also raise a possibility thatRET/PTCrearrangement can be initiated by a complex DSB that is induced in one of the fusion partner genes.