A method for the extraction of nucleic acids from a wide range of environmental samples was developed. This method consists of several modules, which can be individually modified to maximize yields in extractions of DNA and RNA or separations of DNA pools. Modules were designed based on elaborate tests, in which permutations of all nucleic acid extraction steps were compared. The final modular protocol is suitable for extractions from igneous rock, air, water, and sediments. Sediments range from high-biomass, organic rich coastal samples to samples from the most oligotrophic region of the world's oceans and the deepest borehole ever studied by scientific ocean drilling. Extraction yields of DNA and RNA are higher than with widely used commercial kits, indicating an advantage to optimizing extraction procedures to match specific sample characteristics. The ability to separate soluble extracellular DNA pools without cell lysis from intracellular and particle-complexed DNA pools may enable new insights into the cycling and preservation of DNA in environmental samples in the future. A general protocol is outlined, along with recommendations for optimizing this general protocol for specific sample types and research goals.

In marine sediments, DNA occurs both inside and outside living organisms. DNA not enclosed in living cells may account for the largest fraction of total DNA, and include molecules locked within dead cells, organic and inorganic aggregates, adsorbed onto mineral matrices, and viral DNA. This DNA comprises genetic material released in situ from sediment microbial communities, as well as DNA of pelagic and terrestrial origin deposited to the seafloor. DNA not enclosed in living cells undermines the assumption of a direct link between the overall DNA pool and the local, currently living microbial assemblages, in terms of both microbial cell abundance and diversity. At the same time, the extracellular DNA may provide an integrated view of the biodiversity and ecological processes occurring on land, in marine water columns, and sediments themselves, thereby acting as an archive of genetic information which can be used to reconstruct past changes in source environments. In this review, we identify and discuss DNA pools in marine sediments, with special focus on DNA not enclosed in living cells, its origin, dynamics, and ecological and methodological implications. Achievements in deciphering the genetic information held within each DNA pool are presented along with still-standing challenges and major gaps in current knowledge.

Marine sediments harbour extracellular DNA (exDNA) not associated with currently living organisms. Including this exDNA in genetic surveys may distort abundance and diversity estimates of living prokaryotic communities. We separately extract exDNA and intracellular DNA (inDNA) from 11 horizons in a 10-m deep sediment core from Aarhus Bay (Denmark) that spans > 9000 years of Holocene sedimentation. We compare depth profiles of bacterial and archaeal 16S rRNA gene compositions to those of macrofaunal activity (bioturbation), sulfate and methane concentrations, sediment age and lithology. Among these variables, bioturbation shows the strongest relationship with the two DNA pools. In bioturbated surface sediments, the majority of Operational Taxonomic Units (OTUs) present in exDNA is absent from inDNA, thus belonging to microorganisms that were not alive at the time of sampling. Below the bioturbation zone, the two DNA pools display a much higher phylogenetic similarity. At all depths, the majority of exDNA and inDNA sequences show highest sequence similarities to sediment microorganisms, a finding that is additionally supported by separate analyses on low- and high-molecular weight exDNA. Our results indicate that in Aarhus Bay the vast majority of prokaryotic exDNA is turned over, thus not contributing to a genetic archive of past environmental change.

Integrated Ocean Drilling Program Expedition 347 aimed to retrieve sediments from different settings of the Baltic Sea, encompassing the last interglacial-glacial cycle to address scientific questions along four main research themes: 1. Climate and sea level dynamics of marine isotope Stage (MIS) 5, including onsets and terminations; 2. Complexities of the latest glacial, MIS 4-MIS 2; 3. Glacial and Holocene (MIS 2-MIS 1) climate forcing; and 4. Deep biosphere in Baltic Sea Basin (BSB) sediments. These objectives were accomplished by drilling in six subbasins: (1) the gateway of the BSB (Anholt), where we focused on sediments from MIS 6-5 and MIS 2-1; (2) a subbasin in the southwestern BSB (Little Belt) that possibly holds a unique MIS 5 record; (3, 4) two subbasins in the south (Bornholm Basin and Hanö Bay) that may hold long complete records from MIS 4-2; (5) a 450 m deep subbasin in the central Baltic (Landsort Deep) that promises to contain a thick and continuous record of the last ~14,000 y; and (6) a subbasin in the very north (Ångermanälven River estuary) that contains a uniquely varved (annually deposited) sediment record of the last 10,000 y. These six areas were expected to contain sediment sequences representative of the last ~140,000 y, with paleoenvironmental information relevant on a semicontinental scale because the Baltic Sea drains an area four times as large as the basin itself. The location of the BSB in the heartland of a recurrently waning and waxing ice sheet, the Scandinavian Ice Sheet, has resulted in a complex development: repeated glaciations of different magnitudes, sensitive responses to sea level and gateway threshold changes, large shifts in sedimentation patterns, and high sedimentation rates. Its position also makes it a unique link between Eurasian and northwest European terrestrial records. Therefore, the sediments of this largest European intracontinental basin form a rare archive of climate evolution over the latest glacial cycle. High sedimentation rates provide an excellent opportunity to reconstruct climatic variability of global importance at a unique resolution from a marine-brackish setting. Comparable sequences cannot be retrieved anywhere in the surrounding onshore regions. Furthermore, and crucially, the large variability (salinity, climate, sedimentation, and oxygenation) that the BSB has under

Adenosine kinase (AK) catalyzes the phosphorylation of adenosine (Ado) to AMP by means of a kinetic mechanism in which the two substrates Ado and ATP bind the enzyme in a binary and/or ternary complex, with distinct protein conformations. Most of the described inhibitors have Ado-like structural motifs and are nonselective, and some of them (e.g., the tubercidine-like ligands) are characterized by a toxic profile. We have cloned and expressed human AK (hAK) and searched for novel non-substrate-like inhibitors. Our efforts to widen the structural diversity of AK inhibitors led to the identification of novel non-nucleoside, noncompetitive allosteric modulators characterized by a unique molecular scaffold. Among the pyrrolobenzoxa(thia)zepinones (4a-qq) developed, 4a was identified as a non-nucleoside prototype hAK inhibitor. 4a has proapoptotic efficacy, slight inhibition of short-term RNA synthesis, and cytostatic activity on tumor cell lines while showing low cytotoxicity and no significant adverse effects on short-term DNA synthesis in cells.