An Integrated Approach Reveals DNA Damage and Proteotoxic Stress as Main Effects of Proton Radiation in S. cerevisiae

Abstract

Proton radiotherapy (PRT) has the potential to reduce the normal tissue toxicity associated with conventional photon-based radiotherapy (X-ray therapy, XRT) because the active dose can be more directly targeted to a tumor. Although this dosimetric advantage of PRT is well known, the molecular mechanisms affected by PRT remain largely elusive. Here, we combined the molecular toolbox of the eukaryotic model Saccharomyces cerevisiae with a systems biology approach to investigate the physiological effects of PRT compared to XRT. Our data show that the DNA damage response and protein stress response are the major molecular mechanisms activated after both PRT and XRT. However, RNA-Seq revealed that PRT treatment evoked a stronger activation of genes involved in the response to proteotoxic stress, highlighting the molecular differences between PRT and XRT. Moreover, inhibition of the proteasome resulted in decreased survival in combination with PRT compared to XRT, not only further confirming that protons induced a stronger proteotoxic stress response, but also hinting at the potential of using proteasome inhibitors in combination with proton radiotherapy in clinical settings.

Molecular Mechanisms of Resistance to Ionizing Radiation in S. cerevisiae and Its Relationship with Aging, Oxidative Stress, and Antioxidant Activity

“…Exposure to ionizing radiation activates all available cellular machinery to prevent/repair DNA and protein damage [ 12 ]. The repair of DNA damage is mainly carried out by HR and NHEJ.…”

“…In this sense, it is logical that the HSP104 protein is the only common one obtained involved in various metabolic pathways responsible for maintaining longevity and avoiding misfolded proteins. HSP play an important role in cellular proteostasis, also maintaining the correct folding and stabilization of the protein complexes involved in DNA repair and thus contributing to the effectiveness of response mechanisms against DNA-damaging agents [ 12 ]. Krisko and Radman [ 13 ] used HSP104-GFP aggregates as a marker of aging in S. cerevisiae due to the accumulation of misfolded protein aggregates during the yeast lifespan.…”

“…In this sense, the response of proteins to stress caused by radiation also activates molecular response mechanisms, especially chaperones. Heat shock proteins (HSP) play an important role in the maintenance of proteostasis through the correct folding of damaged proteins [ 12 ]. However, when misfolding proteins are oxidized, the damage is fixed and therefore chaperones cannot restore folded proteins correctly.…”

Saccharomyces cerevisiae as a Model System for Eukaryotic Cell Biology, from Cell Cycle Control to DNA Damage Response

“…The integration of DNA barcodes enables identification of individual mutants by their respective barcode, including tracking individual mutants in populations that contain a mix of all 4800 deletion mutants by deep-sequencing the barcode region from bulk samples, and then counting the relative proportion of each barcode [ 14 , 15 , 16 , 17 , 18 ]. The yeast deletion collection was crucial in research assigning specific cellular functions to genes.…”

“…Even now, with the increased toolbox for higher eukaryotes, yeast remains a useful model in radiation biology. For example, to investigate radiation damage and repair after different radiation modalities, such as conventional photon radiotherapy and proton radiotherapy [ 15 , 237 ].…”