Balance of assimilative and dissimilative nitrogen processes in a diatom-rich tidal flat sediment

Tidal flat sediments are subject to repetitive mixing and resuspension events. In a short-term (24 h) <sup>15</sup>N-labelling experiment, we investigated reactive nitrogen cycling in a tidal flat sediment following an experimentally induced resuspension event. We focused on (a) the relative importance of assimilatory versus dissimilatory processes and (b) the role of benthic microalgae therein. <sup>15</sup>N-labelled substrate was added to homogenized sediment, and <sup>15</sup>N was subsequently traced into sediment and dissolved inorganic nitrogen (DIN) pools. Integration of results in a N-cycle model allowed us to quantify the proportion of major assimilatory and dissimilatory processes in the sediment. <br><br> Upon sediment disturbance, rates of dissimilatory processes like nitrification and denitrification were very high, but declined rapidly towards a steady state. Once this was reached, the balance between assimilation and dissimilation in this tidal mudflat was mainly dependent on the nitrogen source: nitrate was utilized almost exclusively dissimilatory via denitrification, whereas ammonium was rapidly assimilated, with about a quarter of this assimilation due to benthic microalgae (BMA). Benthic microalgae significantly affected the nitrogen recycling balance in sediments, because in the absence of BMA activity the recovering sediment turned from a net ammonium sink to a net source. <br><br> The driving mechanisms for assimilation or dissimilation accordingly appear to be ruled to a large extent by external physical forcing, with the entire system being capable of rapid shifts following environmental changes. Assimilatory pathways gain importance under stable conditions, with a substantial contribution of BMA to total assimilation

Nitrification and growth of autotrophic nitrifying bacteria and Thaumarchaeota in the coastal North Sea

Nitrification and the associated growth of autotrophic nitrifiers, as well as the contributions of bacteria and Thaumarchaeota to total autotrophic C-fixation by nitrifiers were investigated in the Dutch coastal North Sea from October 2007 to March 2008. Rates of nitrification were determined by incubation of water samples with ¹⁵N-ammonium and growth of autotrophic nitrifiers was measured by incubation with ¹³C-DIC (dissolved inorganic carbon) in the presence and absence of nitrification inhibitors (nitrapyrin and chlorate) in combination with compound-specific stable isotope (¹³C) analysis of bacterial and Thaumarchaeotal lipid biomarkers. Net nitrification during the sampling period was evident from the concentration dynamics of ammonium, nitrite and nitrate. Measured nitrification rates were high (41–221 nmol N L^-1 h^-1). Ammonium assimilation was always substantially lower than nitrification – with nitrification on average contributing 89% (range 73–97%) to total ammonium consumption.¹³C-DIC fixation into bacterial and Thaumarchaeotal lipids was strongly reduced by the nitrification inhibitors (27–95 %). The inhibitor-sensitive ¹³C-PLFA (phospholipid-derived fatty acid) pool was dominated by the common PLFAs 16:0, 16:1?7c and 18:1?7c throughout the whole sampling period and occasionally also included the polyunsaturated fatty acids 18:2?6c and18:3?3. ¹³C-DIC fixation activity of the nitrifying bacteria was much higher than that of the nitrifying Thaumarchaeota throughout the whole sampling period, even during the peak in Thaumarchaeotal abundance and activity. This suggests that the contribution of autotrophic Thaumarchaeota to nitrification during winter in the coastal North Sea may have been smaller than expected from their gene abundance (16S rRNA and <i>amoA</i> (ammonia monooxygenase)). These results emphasize the importance of direct measurements of the actual activity of bacteria and Thaumarchaeota, rather than abundance measurements only, in order to elucidate their biogeochemical importance. The ratio between rates of nitrification versus DIC fixation by bacterial nitrifiers was higher or even much higher than typical values for autotrophic nitrifiers, indicating that little DIC was fixed relative to the amount of energy that was generated by nitrification

Phospholipid-Derived Fatty Acids and Quinones as Markers for Bacterial Biomass and Community Structure in Marine Sediments

Phospholipid-derived fatty acids (PLFA) and respiratory quinones (RQ) are microbial compounds that have been utilized as biomarkers to quantify bacterial biomass and to characterize microbial community structure in sediments, waters, and soils. While PLFAs have been widely used as quantitative bacterial biomarkers in marine sediments, applications of quinone analysis in marine sediments are very limited. In this study, we investigated the relation between both groups of bacterial biomarkers in a broad range of marine sediments from the intertidal zone to the deep sea. We found a good log-log correlation between concentrations of bacterial PLFA and RQ over several orders of magnitude. This relationship is probably due to metabolic variation in quinone concentrations in bacterial cells in different environments, whereas PLFA concentrations are relatively stable under different conditions. We also found a good agreement in the community structure classifications based on the bacterial PLFAs and RQs. These results strengthen the application of both compounds as quantitative bacterial biomarkers. Moreover, the bacterial PLFA- and RQ profiles revealed a comparable dissimilarity pattern of the sampled sediments, but with a higher level of dissimilarity for the RQs. This means that the quinone method has a higher resolution for resolving differences in bacterial community composition. Combining PLFA and quinone analysis as a complementary method is a good strategy to yield higher resolving power in bacterial community structure

Preclinical pharmacokinetics and metabolism of a novel prototype DNA-PK inhibitor NU7026

In this study we investigated the in vitro time dependence of radiosensitisation, pharmacokinetics and metabolism of NU7026, a novel inhibitor of the DNA repair enzyme DNA-dependent protein kinase (DNA-PK). At a dose of 10 μM, which is nontoxic to cells per se, a minimum NU7026 exposure of 4 h in combination with 3 Gy radiation is required for a significant radiosensitisation effect in CH1 human ovarian cancer cells. Following intravenous administration to mice at 5 mg kg−1, NU7026 underwent rapid plasma clearance (0.108 l h−1) and this was largely attributed to extensive metabolism. Bioavailability following interperitoneal (i.p.) and p.o. administration at 20 mg kg−1 was 20 and 15%, respectively. Investigation of NU7026 metabolism profiles in plasma and urine indicated that the compound undergoes multiple hydroxylations. A glucuronide conjugate of a bis-hydroxylated metabolite represented the major excretion product in urine. Identification of the major oxidation site as C-2 of the morpholine ring was confirmed by the fact that the plasma clearance of NU7107 (an analogue of NU7026 methylated at C-2 and C-6 of the morpholine ring) was four-fold slower than that of NU7026. The pharmacokinetic simulations performed predict that NU7026 will have to be administered four times per day at 100 mg kg−1 i.p. in order to obtain the drug exposure required for radiosensitisation

Collective electrical oscillations of a diatom population induced by dark stress

Diatoms are photosynthetic microalgae, a group with a major environmental role on the planet due to the biogeochemical cycling of silica and global fixation of carbon. However, they can evolve into harmful blooms through a resourceful communication mechanism, not yet fully understood. Here, we demonstrate that a population of diatoms under darkness show quasi-periodic electrical oscillations, or intercellular waves. The origin is paracrine signaling, which is a feedback, or survival, mechanism that counteracts changes in the physicochemical environment. The intracellular messenger is related to Ca2+ ions since spatiotemporal changes in their concentration match the characteristics of the intercellular waves. Our conclusion is supported by using a Ca2+ channel inhibitor. The transport of Ca2+ ions through the membrane to the extracellular medium is blocked and the intercellular waves disappear. The translation of microalgae cooperative signaling paves the way for early detection and prevention of harmful blooms and an extensive range of stress-induced alterations in the aquatic ecosystem.Portuguese Foundation for Science and Technology (FCT) [SFRH/BPD/91518/2012, UID/Multi/04326/2013]; SNMB - INOV: Innovation for a more competitive shellfish sector; Operational Program (OP); European Union through the European Structural Funds and Investment Funds (FEEI); European Maritime and Fisheries Fund (EMFF)info:eu-repo/semantics/publishedVersio

Essential Factors for Incompatible DNA End Joining at Chromosomal DNA Double Strand Breaks In Vivo

Non-homologous end joining (NHEJ) is a major pathway for the repair of DNA double strand break (DSBs) with incompatible DNA ends, which are often generated by ionizing irradiation. In vitro reconstitution studies have indicated that NHEJ of incompatible DNA ends requires not only the core steps of synapsis and ligation, employing KU80/DNA-PKcs and LIG4, but also additional DNA end processing steps, such as DNA end resection by Artemis and gap-filling by POLλ and POLμ. It seems that DNA end processing steps are important for joining of incompatible DNA ends rather than compatible ends. Despite the fact that DNA end processing is important for incompatible DNA end joining in vitro, the role of DNA processing in NHEJ of incompatible DSBs in vivo has not yet been demonstrated. Here we investigated the in vivo roles of proteins implicated in each step of NHEJ using an assay in which NHEJ of incompatible DNA ends on chromosomal DNA can be assessed in living human cells. siRNA- or inhibitor-mediated impairment of factors in each NHEJ step resulted in a reduction in joining efficiency. Strikingly, stronger effects were observed when DNA end resection and ligation protein functions were impaired. Disruption of synapsis by KU80 and DNA-PKcs impairment, or the disruption of gap filling by POLλ and POLμ depletion, resulted in higher levels of microhomology-mediated joining. The present study indicates that DNA end resection and ligation factors are critical for the efficient joining of incompatible ends in vivo, further emphasizing the importance of synapsis and gap-filling factors in preventing illegitimate joining

The BRCT Domain of PARP-1 Is Required for Immunoglobulin Gene Conversion

During affinity maturation, genomic integrity is maintained through specific targeting of DNA mutations. The DNA damage sensor PARP-1 helps determine whether a DNA lesion results in faithful or mutagenic repair