PARENT SESSION

Mutagenesis/Clastogenesis/Carcinogenesis

(PP070) Persistent genomic instability induced in human lymphoblastoid cells exposed to —rays.

Lenarczyk, Marek ^*,1, Lasater, Sarah¹, Jordan, Robert ², Schwartz, Jeffrey ², Liber, Howard ¹, ¹ Department of Environmental and Radiological Health Sciences, Fort Collins, CO, USA² Department of Radiation Oncology, Seattle, WA, USA

ABSTRACT- A number of observations indicate that the induction of persistent genomic instability is an important event in carcinogenesis but most work has been done in cells that have survived high radiation doses. Thus, the objective of our studies was to investigate spontaneous genomic instability as well as instability induced by a low (0.1 Gy) or high (2 Gy) dose of –rays. The studies utilized human lymphoblastoid cells differing in genetic background: i) TK6 cells with wild–type TP53 status, ii) TK6–bcl2 cells, the same as TK6 but with BCL2 over–expression, and iii) TK6–DNA–PKcs cells, the same as TK6 but with transient knockdown of expression of the gene coding for the catalytic subunit of the DNA–dependent protein kinase (DNA–PKcs). Targeted DNA–PKcs gene silencing by RNA interference (RNAi) with small interfering RNAs (siRNAs) was applied. Independent clones from untreated or irradiated TK6, TK6–bcl2 or TK6–DNA–PKcs cells were isolated and examined for evidence of instability at the thymidine kinase (tk) locus more than 35 generations after exposure. In untreated cells, there were no significant differences among cells with different genetic backgrounds. However, irradiation did produce alterations. At the low dose of 0.1 Gy, persistent induced genomic instability was greater in TK6–DNA–PKcs knockdown cells than in TK6. This was true also after 2 Gy. In TK6–bcl2 cells, higher levels of genomic instability also were observed after either the low or high dose exposure; however, only the high dose of 2 Gy gave a statistically significant increase. Based on the above results we hypothesize that: (i) radiation–induced genomic instability in TK6 cells may be partially mediated by apoptosis, and (ii) genomic instability is greater in TK6 cells deficient in DNA double strand break repair by NHEJ at the time of irradiation, and this is true even after a low dose (0.1 Gy) of –rays. Supported by U.S. DOE Grant DE–FG03–02ER63365 (HLL)

Key words: genomic, instability, mutagenesis, cells