The human DNA glycosylases NEIL1 and NEIL3 excise psoralen-induced DNA-DNA cross-links in a four-stranded DNA structure

Interstrand cross-links (ICLs) are highly cytotoxic DNA lesions that block DNA replication and transcription by preventing strand separation. Previously, we demonstrated that the bacterial and human DNA glycosylases Nei and NEIL1 excise unhooked psoralen-derived ICLs in three-stranded DNA via hydrolysis of the glycosidic bond between the crosslinked base and deoxyribose sugar. Furthermore, NEIL3 from Xenopus laevis has been shown to cleave psoralen- and abasic site-induced ICLs in Xenopus egg extracts. Here we report that human NEIL3 cleaves psoralen-induced DNA-DNA cross-links in three-stranded and four-stranded DNA substrates to generate unhooked DNA fragments containing either an abasic site or a psoralen-thymine monoadduct. Furthermore, while Nei and NEIL1 also cleave a psoralen-induced four-stranded DNA substrate to generate two unhooked DNA duplexes with a nick, NEIL3 targets both DNA strands in the ICL without generating single-strand breaks. The DNA substrate specificities of these Nei-like enzymes imply the occurrence of long uninterrupted three- and four-stranded crosslinked DNA-DNA structures that may originate in vivo from DNA replication fork bypass of an ICL. In conclusion, the Nei-like DNA glycosylases unhook psoralen-derived ICLs in various DNA structures via a genuine repair mechanism in which complex DNA lesions can be removed without generation of highly toxic double-strand breaks

DNA glycosylases: in DNA repair and beyond

The base excision repair machinery protects DNA in cells from the damaging effects of oxidation, alkylation, and deamination; it is specialized to fix single-base damage in the form of small chemical modifications. Base modifications can be mutagenic and/or cytotoxic, depending on how they interfere with the template function of the DNA during replication and transcription. DNA glycosylases play a key role in the elimination of such DNA lesions; they recognize and excise damaged bases, thereby initiating a repair process that restores the regular DNA structure with high accuracy. All glycosylases share a common mode of action for damage recognition; they flip bases out of the DNA helix into a selective active site pocket, the architecture of which permits a sensitive detection of even minor base irregularities. Within the past few years, it has become clear that nature has exploited this ability to read the chemical structure of DNA bases for purposes other than canonical DNA repair. DNA glycosylases have been brought into context with molecular processes relating to innate and adaptive immunity as well as to the control of DNA methylation and epigenetic stability. Here, we summarize the key structural and mechanistic features of DNA glycosylases with a special focus on the mammalian enzymes, and then review the evidence for the newly emerging biological functions beyond the protection of genome integrity

Methanogens and Methanogenesis in Hypersaline Environments

Methanogenesis is controlled by redox potential and permanency of anaerobic conditions; and in hypersaline environments, the high concentration of terminal electron acceptors, particularly sulfate, is an important controlling factor. This is because sulfate-reducing microbes, compared with methanogens, have a greater affinity for, and energy yield from, competitive substrates like hydrogen and acetate. However, hypersalinity is not an obstacle to methylotrophic methanogenesis; in many cases hypersaline environments have high concentrations of noncompetitive substrates like methylamines, which derive from compatible solutes such as glycine betaine that is synthesized by many microbes inhabiting hypersaline environments. Also, depletion of sulfate, as may occur in deeper sediments, allows increased methanogenesis. On the other hand, increasing salinity requires methanogens to synthesize or take up more compatible solutes at a significant energetic cost. Acetoclastic and hydrogenotrophic methanogens, with their lower energetic yields, are therefore more susceptible than methylotrophic methanogens, which further explains the predominance of methylotrophic methanogens like Methanohalophilus and Methanohalobium spp. in hypersaline environments. There are often deviations from the picture outlined above, which are sometimes difficult to explain. Identifying the role of uncultivated Euryarchaeota in hypersaline environments, elucidating the effects of different ions (which have differential stress effects and potential as electron acceptors), and understanding the effects of salinity on all microbes involved in methane cycling will help us to understand and predict methane production in hypersaline environments. A good demonstration of this is a recent discovery of extremely haloalkaliphilic methanogens living in hypersaline lakes, which utilize the methyl-reducing pathway and form a novel class “Methanonatronarchaeia” in the Euryarchaeota