Repair of extensive ionizing-radiation DNA damage at 95 degrees C in the hyperthermophilic archaeon Pyrococcus furiosus

Abstract

We investigated the capacity of the hyperthermophile Pyrococcus furiosus for DNA repair by measuring survival at high levels of 60 Co ␥-irradiation. The P. furiosus 2-Mb chromosome was fragmented into pieces ranging from 500 kb to shorter than 30 kb at a dose of 2,500 Gy and was fully restored upon incubation at 95°C. We suggest that recombination repair could be an extremely active repair mechanism in P. furiosus and that it might be an important determinant of survival of hyperthermophiles at high temperatures.Pyrococcus furiosus is a hyperthermophile growing optimally at 100°C. It is a member of the Archaea, a group of prokaryotes which have many molecular features in common with modern eukaryotes, and which are thought by many to be ancestral life forms (25). The exceptional degree of tolerance and resistance of hyperthermophiles to high temperatures must include adaptations that affect all levels of the cellular machinery, including the enzymes that are involved in maintaining the integrity and stability of genomic DNA. Kopylov et al. (14) reported that a closely related member of the Archaea, Thermococcus stetteri, is 12 times more resistant to ␥-irradiation than Escherichia coli but 2 times more sensitive than the bacterium Deinococcus radiodurans. In addition, Peak et al. (20) have shown that at 100°C, the DNA of P. furiosus is 20 times more resistant to thermal breakage in vivo than the DNA from the mesophile E. coli. Thermophilic members of the Archaea have histone proteins (24) known to give partial protection of plasmid DNA to fast neutrons and ␥-photons (12) and to prevent thermal denaturation of DNA (5). However, the protective effect of DNA-binding proteins cannot account for the extreme resistance of these microorganisms to heat and ␥-irradiation (12,14,20), suggesting the presence of very active mechanisms for DNA repair.Among the lesions induced by ionizing radiation in cellular DNA, double-strand breaks (DSBs) are the least efficiently repaired, and their frequency is correlated with cell death (9). Indeed, E. coli and most other organisms cannot survive if more than two or three DSBs are introduced per chromosome, independently of their physiological state (15,21). DSBs are noninformative lesions that affect the DNA double helix at the same site, eliminating intact template for repair and precluding any excision repair processes. However, D. radiodurans has been found to be extremely resistant to ionizing radiation (19). This organism can repair more than 100 DSBs per chromosome, induced by ionizing radiation, without loss of viability (19). Its extreme resistance to ␥-irradiation is attributed to possible adaptation to desiccation (16). RecA-dependent recombinational repair and single-strand annealing are the two mechanisms proposed to account for repair of ionizing-radiation damage in D. radiodurans (16,18).In contrast, there is currently very little information about DNA repair systems in hyperthermophiles, and what information there is consists mainly of the characterization of DNA rep...

Domain Analysis of an Archaeal RadA Protein for the Strand Exchange Activity

“…[1][2][3][4]. Two genes, radA and radB, both of which encode RecA/Rad51 family proteins, have been found in the genomes of archaeal organisms (5,6). All of the euryarchaeotic total genome sequences that have been determined to date contain both RadA and RadB (7,8).…”

Evidence for extensive gene flow and Thermotoga subpopulations in subsurface and marine environments

“…Extensive recombination (repair) has been observed in other hyperthermophilic microorganisms such as Pyrococcus furiosus (Diruggiero et al, 1997), Sulfolobus islandicus (Whitaker et al, 2005) and Persephonella (Mino et al, 2013). Thus, the high levels of recombination observed in Thermotoga might be a by-product of high levels of DNA repair (Johnston et al, 2014).…”

DNA Repair: Lessons from the Evolution of Ionizing- Radiation-Resistant Prokaryotes – Fact and Theory

“…Thus, it is legitimate at this point to assume that the in vivo affinity for DNA of 'any' RecA within D. radiodurans will differ from its in vivo affinity for DNA within an IR-sensitive cell like E. coli. Until more research is done, investigators presently agree that repair of DNA DSBs mediated by recA-like genes is an extremely active and distinct repair mechanism in Deinococcus and Pyrococcus (DiRuggiero et al, 1997;Kim and Cox, 2002;Zahradka et al, 2006;Sghaier et al, 2010). Experimental support for the theory of an ancestral ESDSA repair process is needed.…”

“…I propose the following general definition adopted in a previous paper (Sghaier et al, 2008): An ionizing-radiation-resistant prokaryotes (IRRP) is any vegetative prokaryote that can thrive after exposure to high, acute IR (generally, with a D 10 value -the dose necessary to effect a 90% reduction in Colony Forming Units -greater than 1 kGy) using efficient physiological, genetic and proteic protection and repair mechanisms to fully amend its DNA DSBs. IR resistance has been observed in a broad range of prokaryotic groups (Kopylov et al, 1993), including hyperthermophilic Archaea (P. abyssi, P. furiosus, Thermococcus marinus, Thermococcus radiotolerans and Thermococcus gammatolerans) (DiRuggiero et al, 1997;Jolivet et al, 2003a;Jolivet et al, 2003b;Jolivet et al, 2004), halophilic Archaea (Halobacterium sp.) (Kottemann et al, 2005), the Deinococcus-Thermus group (many Deinococcus sp.…”

“…The PolA-RecAmediated repair process possesses the following important characteristics: 1. Functionally, its proteins (PolA and RecA) are important for IR resistance (DiRuggiero et al, 1997;Cox and Battista, 2005;Zahradka et al, 2006;Galhardo and Rosenberg, 2009;Slade et al, 2009;Repar et al, 2010). 2.…”