Ⅲ-4. Radiation Chemistry & Biology

RIKEN Accel. Prog. Rep. 47 (2014)

Effects of trichostatin A on radiosensitivity to high-linear energy transger(LET) radiation in mammalian cells with defects in DNA repair proteins
M. Izumi*1 and T. Abe*1
In eukaryotes, DNA is associated with histones andpackaged into nucleosomes, which are arranged into higher
order structures to form chromatin.
The chromatin
structure contributes to various aspects of DNA metabolism
including replication, recombination, and transcription.
However, it is still unclear how repair reactions andcheckpoint responses caused by heavy-ion irradiation areregulated by chromatin structures. To investigate the rolesof chromatin structures in DNA repair after heavy-ionirradiation, we have been focusing on the damage response
observed after cells are treated with a potent histone
deacetylase inhibitor, trichostatin A (TSA).
To analyze the effects of TSA on repair pathways, weinvestigated the X-ray sensitivity of wild-type CHO-AA8
cells and CHO mutant lines deficient in homologous
recombination (irs1SF cells) 1), non-homologousend-joining (V3 cells) 2), and base excision repair (EM9cells) 3) in the absence or presence of TSA in a previousstudy4). All three mutant cell lines showed increased
X-ray sensitivity (Fig. 1a). TSA treatment enhanced the
X-ray sensitivity of wild-type CHO cells. In contrast, TSA
enhanced the X-ray radioresistance of irs1SF cells,suggesting that the homologous recombination pathway is
involved in radiosensitivity enhancement by TSA.
However, TSA did not affect V3 and EM9 survival. Theseresults suggest that non-homologous end-joining and/orbase excision repair are stimulated by TSA.
In this study, we investigated argon-ion (LET = 300keV/µm) sensitivity using the same cell lines. We
compared the radiosensitivity of the four cell lines without
TSA treatment (Fig. 1a). We found that CHO and V3 cells
showed nearly identical dose-response profiles afterargon-ion irradiation. We obtained the same results after
carbon-ion (LET = 80 keV/µm) irradiation (data not shown),suggesting that non-homologous end-joining is not involvedin the repair pathway induced by high-LET ionizing
radiation.
EM9 and irs1SF cells showed increased
sensitivity to argon ions. These results are compatible
with those of several recent studies5,6).
We investigated the effect of TSA on cell line
survival (Fig. 1b). TSA treatment enhanced CHO cellsensitivity to argon ions. However, TSA did not affectirs1SF cell sensitivity, which is compatible with theconclusion that non-homologous end-joining is not involvedin the repair pathway induced by high-LET irradiation. Incontrast, TSA slightly enhanced V3 and EM9 cellradiosensitivity, which seems to be due to the inhibitory
effect of TSA on homologous recombination.
Although our survival assay suggests that thenon-homologous pathway is not involved in repair after
heavy-ion irradiation, we observed that DNA-PK wasrecruited to the DNA damage sites (data not shown).
Currently, we are investigating the localization of repair
proteins by indirect immunofluorescence and studying themechanism underlying the suppression of non-homologous
end-joining after heavy-ion irradiation.

Fig. 1 (a) X-ray and argon-ion sensitivity of CHO, irs1SF,V3, and EM9 cells. Cells were irradiated with X-rays orargon ions (LET = 300 keV/µm), and radiosensitivity wasestimated by the clonogenic survival assay. (b) Effects oftrichostatin A (TSA) on the radiosensitivity of CHO, irs1SF,
V3, and EM9 cells. Cells were pretreated with TSA (0.1µM) for 10 h and irradiated with argon ions. Subsequently,
the cells were cultured for an additional 14 h in the presenceof TSA, and radiosensitivity was estimated by theclonogenic survival assay. Abbreviations: wt, wild type;HR, homologous recombination; NHEJ, non-homologous
end-joining; BER, base excision repair.
References
1) R.S. Tebbs, et al.: Proc. Natl. Acad. Sci. USA 92, 6354 (1995)
2) S.R. Peterson, et al.: Proc. Natl. Acad. Sci. USA 92, 3171
(1995)
3) L.H. Thompson, et al.: Mol. Cell. Biol. 10, 6160 (1990)
4) M. Izumi, et al.: RIKEN Accel. Prog. Rep. 46, 254 (2013)5) H. Wang, et al: Nucl. Acids Res. 38, 3245 (2010)6) S.C. Genet, et al.: Oncology Rep. 28, 1591 (2012)*1 RIKEN Nishina Center- 283 -