Repair of DNA Double-Strand Breaks following UV Damage in ThreeSulfolobussolfataricusStrains

Abstract

DNA damage repair mechanisms have been most thoroughly explored in the eubacterial and eukaryotic branches of life. The methods by which members of the archaeal branch repair DNA are significantly less well understood but have been gaining increasing attention. In particular, the approaches employed by hyperthermophilic archaea have been a general source of interest, since these organisms thrive under conditions that likely lead to constant chromosomal damage. In this work we have characterized the responses of three Sulfolobus solfataricus strains to UV-C irradiation, which often results in double-strand break formation. We examined S. solfataricus strain P2 obtained from two different sources and S. solfataricus strain 98/2, a popular strain for site-directed mutation by homologous recombination. Cellular recovery, as determined by survival curves and the ability to return to growth after irradiation, was found to be strain specific and differed depending on the dose applied. Chromosomal damage was directly visualized using pulsed-field gel electrophoresis and demonstrated repair rate variations among the strains following UV-C irradiation-induced double-strand breaks. Several genes involved in double-strand break repair were found to be significantly upregulated after UV-C irradiation. Transcript abundance levels and temporal expression patterns for doublestrand break repair genes were also distinct for each strain, indicating that these Sulfolobus solfataricus strains have differential responses to UV-C-induced DNA double-strand break damage.Cells have evolved molecular mechanisms to meet the challenge of maintaining genomic integrity by rapidly responding to environmental stresses that can damage proteins and DNA. One of the most common forms of damage is caused by UV light (UV) exposure. High-energy short-wavelength UV-C light is absorbed directly by DNA and induces both cyclobutane pyrimidine dimers between adjacent thymidine or cytosine residues as well as pyrimidine-pyrimidone photoproducts between adjacent pyrimidine residues. Mechanisms for repair of these lesions appear to be present in all organisms and are thought to occur through either light-independent nucleotide excision repair (NER) or light-dependent photoreactivation using photolyases (for reviews, see references 33 and 34). UV-C irradiation also causes the production of reactive oxygen species, which can result in DNA double-strand breaks (DSBs) (6, 44). Our primary understanding for repair of DSBs has come from studies focused primarily on bacteria and eukaryotes. In Escherichia coli, these breaks are repaired through the action of the RecA protein, which assists in recombinational repair of single-strand regions produced through replication fork arrest at UV lesions and in DSB repair by extended synthesis-dependent strand annealing (SDSA) and homologous recombination (9, 19). Eukaryotes employ nonhomologous end joining as well as DSB repair by SDSA and homologous recombination mechanisms to repair these breaks (for recent reviews, see ref...

Biochemical and Functional Characterization of the NurA-HerA Complex from Deinococcus radiodurans

“…In archaea, both nurA and herA are upregulated after gamma irradiation treatment, indicating that these genes might contribute to a DNA DSB repair process (19,20). We found that the two genes were slightly upregulated after gamma irradiation treatment of D. radiodurans cells (data not shown); however, no significant reductions in gamma irradiation or UV irradiation resistance were observed in nurA-, herA-, or nurA-herA-disrupted strains (see Fig.…”

“…The nurA gene encodes a 5=-to-3= ssDNA/dsDNA exonuclease/endonuclease (15), and herA encodes an ATP-dependent bipolar helicase that unwinds dsDNA in both the 5=-to-3= and 3=-to-5= directions (16,17). Usually, mre11-rad50 colocalizes with this operon (18), and nurA, herA, mre11, and rad50 are cotranscribed in response to UV irradiation (19,20). Functional interactions between NurA-HerA and Mre11-Rad50 indicate that these four proteins are involved in HR DNA end resection (13,21).…”

Protein splicing of a recombinase intein induced by ssDNA and DNA damage

“…Although DNA damage has been shown to induce RadA expression in most archaea (Rolfsmeier et al 2010), in the close Pho relative Pyrococcus furiosus, RadA is not induced by several DNA-damaging agents, suggesting a post-translational regulatory mechanism to control activity (Komori et al 2000b). It is unknown whether Pho shares this trait with P. furiosus, but, in any event, intein splicing in response to ssDNA represents an immediate mechanism to post-translationally activate RadA, providing the cell a much faster avenue than de novo transcription and translation to produce functional RadA under stress in response to ssDNA.…”

Effect of Amalaki rasayana on DNA damage and repair in randomized aged human individuals

“…UVC can also induce reactive oxygen species that results in double strand breaks (DSBs). Further, it is known that UVC light arrests DNA replication which may be related to the formation of DSBs at stalled replication forks (Rolfsmeier et al, 2010). It has also been reported that UVC induces significantly more DSBs and chromosomal aberrations in NER-deficient cells indicating conversion of primary non-repaired UVC light induced DNA lesions into DSBs (Dunkern and Kaina, 2002).…”