Protein damage and death by radiation in Escherichia coli and Deinococcus radiodurans

Abstract

Deinococcus radiodurans is among a small number of bacterial species that are extremely resistant to ionizing radiation, UV light, toxic chemicals, and desiccation. We measured proteome oxidation (i.e., protein carbonylation, PC) in D. radiodurans as well as in standard and evolved resistant strains of Escherichia coli exposed to ionizing radiation or UVC light and found a consistent correlation with cell killing. The unique quantitative relationship between incurred PC and cell death holds over the entire range of killing for all tested bacteria and for both lethal agents, meaning that both bacterial species are equally sensitive to PC. We show that the extraordinary robustness of D. radiodurans depends on efficient proteome protection (but not DNA protection) against constitutive and radiation-induced PC consisting of low molecular weight cytosolic compounds. Remarkably, experimental evolution of resistance to ionizing radiation in E. coli coevolves with protection against PC. The decline in biosynthetic efficacy of the cellular proteome, as measured by the loss of reproduction of undamaged bacteriophage lambda in irradiated standard and evolved ionizing radiation-resistant E. coli, correlates with radiation-induced oxidative damage to host cells and their sensitivity to ionizing radiation. This correlation suggests that cell death by radiation is caused primarily by oxidative damage with consequential loss of maintenance activities including DNA repair.

Fig. 1.

Cell death induced by ionizing and UV radiation correlates with ROS production and protein carbonylation in E. coli (Eco) and D. radiodurans (Dra). (A) Cell survival (cfu) and PC versus γ radiation dose. (B) Cell survival and PC versus UVC radiation dose. (C) Intracellular ROS production by γ radiation measured by intracellular DHR in two bacterial species. (D) Intracellular ROS production by UVC radiation measured by intracellular DHR in two bacterial species. Cell survival and PC are shown as the mean and SD of two duplicate experiments (see also Fig. S1).

Fig. 2.

Single correlation between cell killing and PC. A similar correlation is observed for the two bacterial species, including the resistant E. coli strains CB1000 and CB2000, and for both means of irradiation. Cell survival and PC are shown as the mean and SD of two duplicate experiments.

Fig. 3.

Correlations between radiation-induced protein carbonylation (▲), cellular capacity to produce phage λ (○), and cell death (●) in E. coli. (A) UVC radiation. (B) γ-radiation. Host cell survival (cfu), the cellular capacity to produce phage (infective centers, IC) and PC are presented as the mean and SD of two duplicate experiments (see also Fig. S2).

Fig. 4.

Cell death and protein carbonylation induced by (A) ionizing and (B) UVC radiation in ΔrecA mutants of D. radiodurans. (A) Cell survival (cfu) and total protein carbonylation versus γ irradiation dose. (B) Cell survival (cfu) and total protein carbonylation versus UVC irradiation dose. Cell survival and PC are shown as the mean and SD of two duplicate experiments.