Simultaneous induction of dispersed and clustered DNA lesions compromises DNA damage response in human peripheral blood lymphocytes

Lei Cheng, Beata Brzozowska, Alice Sollazzo, Lovisa Lundholm, Halina Lisowska, Siamak Haghdoost, Andrzej Wojcik

Published: October 31, 2018
https://doi.org/10.1371/journal.pone.0204068

Abstract
Due to its ability to induce DNA damage in a space and time controlled manner, ionising radiation is a unique tool for studying the mechanisms of DNA repair. The biological effec- tiveness of ionising radiation is related to the ionisation density which is defined by the linear energy transfer (LET). Alpha particles are characterised by high LET, while X-rays by low LET values. An interesting question is how cells react when exposed to a mixed beam of high and low LET radiation.
In an earlier study carried out with human peripheral blood lym- phocytes (PBL) we could demonstrate that alpha radiation X-rays interact in producing more chromosomal aberrations than expected based on additivity. The aim of the present investi- gation was to look at the mechanism of the interaction, especially with respect to the ques- tion if it is due to an augmented level of initial damage or impaired DNA repair. PBL were exposed to various doses of alpha particles, X-rays and mixed beams. DNA damage and the kinetics of damage repair was quantified by the alkaline comet
assay. The levels of phosphorylated, key DNA damage response (DDR) proteins ATM, p53 and DNA-PK were measured by Western blotting and mRNA levels of 6 damage-responsive genes were mea- sured by qPCR. Alpha particles and X-rays interact in inducing DNA damage above the level predicted by assuming additivity and that the repair of damage occurs with a delay. The activation levels of DDR proteins and mRNA levels of the studied genes were highest in cells exposed to mixed beams. The results substantiate the idea that exposure to mixed beams presents a challenge for the cellular DDR system.