Short communicationPredicted ionisation in mitochondria and observed acute changes in the mitochondrial transcriptome after gamma irradiation: A Monte Carlo simulation and quantitative PCR study
Highlights
► Investigate radiation effects in cytoplasm, cell nucleus and mitochondria. ► Simulations suggest preferential localization of ionization events in mitochondria. ► qPCR results confirm acute, differential mitochondrial gene expression changes. ► Suggest to incorporate mitochondrial response when estimating radiation effects.
Introduction
Most studies of cellular responses to ionising radiation are centred on the nuclear DNA, whereby the DNA repair processes, rather than the damage directly, are used as proxy read-outs to determine the extent of nuclear DNA damage (Aziz et al., 2012). However, significant effects of ionising radiation on mitochondrial functions (Hwang et al., 1999, Yukawa et al., 1985), mitochondrial oxidative stress (Hosoki et al., 2012, Kobashigawa et al., 2011, Motoori et al., 2001, Tulard et al., 2003) and apoptotic pathways (Belka et al., 2000, Chen et al., 2003, Leach et al., 2001, Zhao et al., 1999) have been reported. Indeed, some experimental observations indicate that the mitochondrial genome may be more susceptible to damaging effects by gamma irradiation than the nuclear genome (Gong et al., 1998, May and Bohr, 2000, Morales et al., 1998), possibly by virtue of the greater likelihood of mitochondria suffering oxidative damage (Yakes and Houten, 1997). In addition to direct radiation effects on mitochondria, mitochondria dysfunction may exert an indirect influence on the nucleus and perpetuate radiation-induced genomic instability (Kim et al., 2006a, Kim et al., 2006b, Miller et al., 2008).
Monte Carlo track structure simulations (Zaider et al., 1983) can be used to estimate likely regions of radiation damage within the cell (Alard et al., 2002, Chauvie et al., 2007, Miller et al., 2000). To date, however, track structure simulations have mostly focused on predicting the occurrence of single or double strand breaks in nuclear DNA as a result of physical processes leading to ionisation formation (Grosswendt, 2005, Nikjoo and Goodhead, 1991, Nikjoo et al., 1999). Here, we employ Monte Carlo simulation and develop a more realistic cell model containing both cell nucleus and mitochondria, as well as currently available data on the elemental concentration in mitochondria (Ernster and Lindberg, 1958, Taylor et al., 1999), in order to predict regions mostly likely to be subject to damage from ionisation formation. We base our model for the simulation on the human leukemic JURKAT cell line, which has previously been used to investigate radiation effects in vitro (Cataldi et al., 2009, Syljuasen and McBride, 1999, Vigorito et al., 1999). In suspension, the cells are of near-spherical shape (Rosenbluth et al., 2006, Roskams and Rodgers, 2002) and are well characterised with respect to the relative volume taken up by mitochondria (Cataldi et al., 2009, Chaigne-Delalande et al., 2008, Kawahara et al., 1998, Ueda et al., 1998, Yasuhara et al., 2003), and their geometry can, therefore, be modelled relatively easily.
PCR or quantitative PCR (qPCR) has previously been used to measure changes in gene expression levels after radiation exposure (Gong et al., 1998, Gubina et al., 2010, Kulkarni et al., 2010). To validate in vitro the predictions made by our Monte Carlo simulation that mitochondria respond to radiation exposure, we quantify by qPCR in JURKAT cells the acute changes in the expression levels of mitochondrial electron transport chain genes as well as mitochondrial transfer RNAs and ribosomal RNAs, in response to a single radiation dose ranging from 10 to 100 Gy.
In the present study, we introduce for the first time a model that specifically includes the mitochondrial compartment, and makes realistic assumptions in regard to the content of those atomic elements in mitochondria that are important for predicting the likely localisation of ionisation events. The qPCR results provide biological evidence that mitochondria are involved in the early cell response to gamma radiation.
Section snippets
Monte Carlo simulation
Simulations were performed using the open-source, general-purpose Monte Carlo radiation transport simulation toolkit Geant4 version 9.4.p02 (Agostinelli et al., 2003, Allison et al., 2006).
Energy deposition and ionisation events in nucleus, cytoplasm and mitochondria
Using a cell model based on the idealised geometry of JURKAT cells (Fig. 1), we performed Monte Carlo radiative transport simulations to examine the dose received as well as the total number of ionisations formed in different compartments of the cell (nucleus, cytoplasm, mitochondria). Fig. 2 shows the dose and ionisation number for each organelle in the left and right panels, respectively.
A higher mitochondrial density increases the overall dose received by each cellular component considered.
Conclusions
Our predicted and observed results draw attention to the importance of mitochondria as a direct target that is likely to influence the immediate response to radiation. Current models do not specifically take into account any possibility of mitochondrially mediated post-radiation effects. However, for a more comprehensive understanding of the overall cellular radiation effects, not only the changes in the nuclear but also the mitochondrial genome is important (Schilling-Toth et al., 2011). Apart
Acknowledgements
Special thanks should be given to Mr Sohil Sheth and Mr Allan Perry, for their technical assistance in performing the irradiation experiments. Dr Cy Jeffries, for his suggestions on the experimental design and critical comments on this manuscript. Dr Alessandra Malaroda, for her critical reading and discussion of the manuscript. Mrs Geetanjali Dhand, for her generous assistance in improving the image quality.
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These authors contributed equally to this work.