Variation in chromosomal aberration induction in differing ATM and ASPM genotypic backgrounds detected via a modified PNA FISH technique

Abstract

A modified PNA (peptide nucleic acid) FISH (fluorescence in situ hybridization) method has been developed for efficient quantitative and qualitative analysis of chromosomal aberration formation. PNA FISH was used to detect a heightened sensitivity in mouse fibroblasts in response to ionizing radiation with Atm-/- and in mouse embryonic fibroblasts (MEFs) in response to loss of Aspm gene respectively. For Atm analysis, four commonly used inbred mouse strains were used, C57BL/6J, A/J, 129S6, and BALB/cByJ, and exposed to cesium-137 at a dose rate of approximately 2.5 Gy/min. For Aspm analysis, MEF cell lines were used and exposed to 0.2 μM tamoxifen for inducible Cre-lox excision of floxed Aspm alleles. Upon analysis of chromosome-type aberrations, there were statistically significant elevations in acentric fragment and/or dicentric average frequencies seen across all strain backgrounds of the Atm-/- genotype, however, only the 129S6 Atm-/- strain backgrounds exhibited a significant increase in mean telomere-associated fusion events. In addition, there was a heightened dose response from a 0 to 2 Gy dose in Atm-/- BALB/cByJ and C57BL/6J backgrounds indicating a lack of synergism of the Prkdc (protein kinase, DNA-activated, catalytic polypeptide) defect in BALB/cByJ strains. For Aspm analysis, in comparison to control, or untreated, MEF strains, there was an increased average aberration frequency in Aspm-/- cell lines over a 48-hour time period. However, there was also an increased frequency in Aspm+/- cell lines comparable to the frequency observed in Aspm-/- cells, which is characteristic of haploinsufficiency. Therefore, not only is Atm status an important determinant of radiosensitivity, but strain backgrounds can also contribute to a heightened response. In addition, Aspm expression is critical for the maintenance and repair of DNA. Together, this data suggests that the Atm and Aspm genes are crucial components in the preservation of chromosomal integrity and genomic stability in response to DNA damage.