DOE Low Dose Radiation Research Program Workshop I Abstracts November 10-12, 1999, Washington, D.C.

14. Genomic Instability Induced by Low Dose Irradiation

Helen H. Evans, Martina L. Veigl, and W. David Sedwick
Department of Radiation Oncology, Case Western Reserve University School of Medicine (BRB), 2109 Adelbert Road, Cleveland, OH 44106-4942
hhe@po.cwru.edu

Summary: The induction of genomic instability by low doses of ionizing radiation will be determined by measuring the delayed increase in mutation frequency in a target gene that requires a frameshift mutation for the easily and directly quantifiable expression of the green fluorescent protein.

Abstract: The objective of the research described in this application is to determine the dose response of the induction of genomic instability by low doses of ionizing radiation, as indicated by a delayed increase in mutation frequency. The delayed increase in mutant frequency will be measured in stable human colon carcinoma cell lines transfected with a vector carrying a reporter green fluorescent protein (GFP) that requires a frame shift mutation for activation. Expression of GFP results in quantifiable colored cells. Because the delayed increase in mutant frequency caused by the induction of genomic instability can be visualized and quantified directly in cells surviving irradiation, it can be detected quickly among many thousands of transfected cells. Such a strategy is necessary for the quantification and investigation of the induction of genomic instability by low doses of ionizing radiation. In addition, the approach can facilitate distinction between the immediate and delayed effects of exposure to ionizing radiation. The induction of genomic instability is relevant with regard to risk estimation since it is thought to be a necessary step in the carcinogenic process.

The hypothesis driving the research is that the induction of genomic instability leading to an delayed increase in the frequency of gene mutations many generations after exposure to radiation can be initiated by radiation-induced, complex, poorly repaired DNA damage that leads to a mutator phenotype. We aim to (1) measure the dose response of the induction of genomic instability by Cs gamma radiation with doses ranging from 0.05 Gy to 3 Gy in order to determine if there is a dose-dependent induction of genomic instability and if there is a threshold apparent in the dose response curve; (2) measure the kinetics of the induction and termination of genomic instability at intervals following exposure to irradiation; (3) compare the induction of genomic instability caused by hydrogen peroxide with that caused by radiation to determine whether or not clustered lesions are required for the induction of genomic instability; (4) measure the dose-response of the induction of genomic instability in cell lines varying in their genetic background with regard to mismatch repair capacity, and the activity of the p53 and p21 genes; and (5) isolate and characterize the unstable cell lines in order to investigate the initial lesion inducing the instability.

The characterization of the induction by low doses of radiation of genomic instability, a process that is thought to be necessary for the carcinogenic process, will be helpful in the estimation of the risk of cancer in persons exposed to these low radiation doses.