Reconstructing the chronology of the natural and anthropogenic uranium isotopic signals in a marine sediment core from Beppu Bay, Japan.

The long-lived U isotopes, 233U and 236U, have been used increasingly in recent years as marine circulation tracers and for identifying sources of uranium contamination in the environment. The sedimentation histories of these two U isotopes in combination with natural 238U were reconstructed for an anoxic sediment core collected from Beppu Bay, Japan, in the western North Pacific Ocean showing good time resolution (less than 2.6 y/sample). The 233U/236U atom ratio showed a prominent peak of (3.20 ± 0.30) × 10-2 around 1957 which can be attributed to the input from atmospheric nuclear weapons tests including thermonuclear tests conducting in the Equatorial Pacific. The integrated 233U/236U ratio of (1.64 ± 0.08) × 10-2 for the sediment was found to be in relatively good agreement with the representative ratio published for global fallout (∼1.4 × 10-2). A prominent increase in the authigenic ratio of 233U/238Ua,s in the leached fraction (1.39 ± 0.11 × 10-11) and the bulk digestion (1.36 ± 0.10 × 10-11) was also observed around 1957. This reflects the input supply of 233U to the seawater which is known to have a relatively constant 238U content. The authigenic 236U/238Ua,s ratio (0.18 ± 0.02 × 10-9) obtained for 1921 increased from the early 1950's to a maximum of (6.59 ± 0.60) × 10-9 around 1962. The variation in this ratio represents well the introduction history of U into the surface environment without site-specific U contamination and the time profile is also consistent with the 137Cs signature. This work thus provides a benchmark for the long-term use of the isotopic U composition as an input parameter for seawater circulation tracers and as a chronological marker for anoxic sediments and sedimentary rocks. Especially the 233U/236U ratio may serve as a key-marker for the new geological age Anthropocene.

Deciphering sources of U contamination using isotope ratio signatures in the Loire River sediments: Exploring the relevance of 233U/236U and stable Pb isotope ratios.

A broad range of contaminants has been recorded in sediments of the Loire River over the last century. Among a variety of anthropogenic activities of this nuclearized watershed, extraction of uranium and associated activities during more than 50 years as well as operation of several nuclear power plants led to industrial discharges, which could persist for decades in sedimentary archives of the Loire River. Highlighting and identifying the origin of radionuclides that transited during the last decades and were recorded in the sediments is challenging due to i) the low concentrations which are often close or below the detection limits of routine environmental surveys and ii) the mixing of different sources. The determination of the sources of anthropogenic radioactivity was performed using multi-isotopic fingerprints (236U/238U, 206Pb/207Pb and 208Pb/207Pb) and the newly developed 233U/236U tracer. For the first time 233U/236U data in a well-dated river sediment core in the French river Loire are reported here. Results highlight potential sources of contamination among which a clear signature of anthropogenic inputs related to two accidents of a former NUGG NPP that occurred in 1969 and 1980. The 233U and 236U isotopes were measured by recent high performance analytical methods due to their ultra-trace levels in the samples and show a negligible radiological impact on health and on the environment. The determination of mining activities by the use of stable Pb isotopes is still challenging probably owing to the limited dissemination of the Pb-bearing material marked by the U-ore signature downstream to the former U mines.

70-Year Anthropogenic Uranium Imprints of Nuclear Activities in Baltic Sea Sediments.

A strongly stratified water structure and a densely populated catchment make the Baltic Sea one of the most polluted seas. Understanding its circulation pattern and time scale is essential to predict the dynamics of hypoxia, eutrophication, and pollutants. Anthropogenic 236U and 233U have been demonstrated as excellent transient tracers in oceanic studies, but unclear input history and inadequate long-term monitoring records limit their application in the Baltic Sea. From two dated Baltic sediment cores, we obtained high-resolution records of anthropogenic uranium imprints originating from three major human nuclear activities throughout the Atomic Era. Using the novel 233U/236U signature, we distinguished and quantified 236U inputs from global fallout (45.4-52.1%), Chernobyl accident (0.3-1.8%), and discharges from civil nuclear industries (46.1-54.3%) to the Baltic Sea. We estimated the total release of 233U (7-15 kg) from the atmospheric nuclear weapon testing and pinpointed the 233U peak signal in the mid-to-late 1950s as a potential time marker for the onset of the Anthropocene Epoch. This work also provides fundamental 236U data on Chernobyl accident and early discharges from civil nuclear facilities, prompting worldwide 233U-236U tracer studies. We anticipate our data to be used in a broader application in model-observation interdisciplinary research on water circulation and pollutant dynamics in the Baltic Sea.

An unknown source of reactor radionuclides in the Baltic Sea revealed by multi-isotope fingerprints.

We present an application of multi-isotopic fingerprints (i.e., 236U/238U, 233U/236U, 236U/129I and 129I/127I) for the discovery of previously unrecognized sources of anthropogenic radioactivity. Our data indicate a source of reactor 236U in the Baltic Sea in addition to inputs from the two European reprocessing plants and global fallout. This additional reactor 236U may come from unreported discharges from Swedish nuclear research facilities as supported by high 236U levels in sediment nearby Studsvik, or from accidental leakages of spent nuclear fuel disposed on the Baltic seafloor, either reported or unreported. Such leakages would indicate problems with the radiological safety of seafloor disposal, and may be accompanied by releases of other radionuclides. The results demonstrate the high sensitivity of multi-isotopic tracer systems, especially the 233U/236U signature, to distinguish environmental emissions of unrevealed radioactive releases for nuclear safeguards, emergency preparedness and environmental tracer studies.

On the Quality Control for the Determination of Ultratrace-Level 236U and 233U in Environmental Samples by Accelerator Mass Spectrometry.

Our recent attempt to determine ultratrace-level 236U and 233U in small-volume seawater samples was challenged by high and unstable procedure blanks in our environmental radioactivity laboratory, which used to be a spent fuel research facility. Through intercomparison experiments with different laboratories and background checks on the chemical reagents and laboratory dust, the resuspended U-bearing dust was identified as the dominating source of the 236U and 233U contamination. With the implementation of background control (especially dust control) measures, the procedure blanks and detection limits of 236U and 233U for the radiochemical separation procedure have been significantly improved by three orders of magnitudes. With well-controlled blanks, the analytical precision for 236U and 233U predominantly relies on the AMS counting statistics. Background check and dust control are strongly recommended before the analyses of environmental-level long-lived radionuclides (such as 236U and 233U) that are conducted in the former or active nuclear facilities, even if clearance of radioactivity relevant for radioprotection was achieved.

First dataset of 236U and 233U around the Greenland coast: A 5-year snapshot (2012-2016).

We report the first combined dataset of 236U and 233U in the Greenland marine environment during the period of 2012-2016. Results are discussed in terms of time evolution and spatial distribution of 236U concentration, and atomic ratios of 236U/238U and 233U/236U. 236U concentrations along the Greenland coast are distributed within a relatively narrow range of (0.7-12.9) × 107 atom/L, corresponding to 236U/238U atomic ratios of (1.1-15.5) × 10-9. The 233U/236U atomic ratios obtained vary from 0.12 × 10-2 to 1.16 × 10-2, with the majority distributed in the range of (0.2-0.7) × 10-2. We applied 233U/236U and 236U/238U atomic ratios in a binary mixing model to identify possible 236U source terms. The results indicate that anthropogenic 236U and 233U in Greenland surface seawater originated from the direct global fallout (DGF) and the Sellafield and La Hague reprocessing plants (RP) is diluted by a third endmember, mostly likely natural ocean water (NOW), containing marginal 236U and 233U. A preliminary estimation of the source terms of 236U using 233U/236U atomic ratios indicate that, for both eastern and western Greenland seawater, contributions from global fallout (GF) constitute about 30% of 236U. The dominating source for 236U, i.e. 70 %, is associated to reactor 236U including discharges from RP and local reactor input in the Arctic Ocean.

Ultratrace Determination of 99Tc in Small Natural Water Samples by Accelerator Mass Spectrometry with the Gas-Filled Analyzing Magnet System.

In the frame of studies on the safe disposal of nuclear waste, there is a great interest for understanding the migration behavior of 99Tc. 99Tc originating from nuclear energy production and global fallout shows environmental levels down to 107 atoms/g of soil (∼2 fg/g). Extremely low concentrations are also expected in groundwater after diffusion of 99Tc through the bentonite constituting the technical barrier for nuclear waste disposal. The main limitation to the sensitivity of the mass spectrometric analysis of 99Tc is the background of its stable isobar 99Ru. For ultratrace analysis, the Accelerator Mass Spectrometry (AMS) setup of the Technical University of Munich using a Gas-Filled Analyzing Magnet System (GAMS) and a 14 MV Tandem accelerator is greatly effective in suppressing this interference. In the present study, the GAMS setup is used for the analysis of 99Tc in samples of the seawater reference material IAEA-443, a peat bog lake, and groundwater from an experiment of in situ diffusion through bentonite in the controlled zone of the Grimsel Test Site (GTS) within the Colloid Formation and Migration (CFM) project. With an adapted chemical preparation procedure, measurements of 99Tc concentrations at the fg/g levels with a sensitivity down to 0.5 fg are accomplished in notably small natural water samples. The access to these low concentration levels allows for the long-term monitoring of in situ tracer tests over several years and for the determination of environmental levels of 99Tc in small samples.

Assessment of 53Mn deposition on Earth via accelerator mass spectrometry.

The 53Mn flux onto Earth is a quantity relevant for different extraterrestrial and astrophysical questions. It is a proxy for related fluxes, such as supernova-produced material or interplanetary dust particles. In this work, we performed a first attempt to assess the 53Mn flux by measuring the 53Mn/10Be isotopic ratio in a 1400 L sample of molten Antarctic snow by AMS (Accelerator Mass Spectrometry). Using the 10Be production rate in the atmosphere, an upper limit of 5.5 × 103 atoms cm-2 yr-1 was estimated for the deposition of extraterrestrial 53Mn. This result is compatible with one of the two discrepant values existing in the literature.

Plutonium Isotopes (239-241Pu) Dissolved in Pacific Ocean Waters Detected by Accelerator Mass Spectrometry: No Effects of the Fukushima Accident Observed.

The concentration of plutonium (Pu) and the isotopic ratios of 240Pu to 239Pu and 241Pu to 239Pu were determined by accelerator mass spectrometry (AMS) in Pacific Ocean water samples (20 L each) collected in late 2012. The isotopic Pu ratios are important indicators of different contamination sources and were used to identify a possible release of Pu into the ocean by the Fukushima Daiichi Nuclear Power Plant (FDNPP) accident. In particular, 241Pu is a well-suited indicator for a recent entry of Pu because 241Pu from fallout of nuclear weapon testings has already significantly decayed. A total of 10 ocean water samples were prepared at the Radiochemie München of the TUM and analyzed at the Vienna Environmental Research Laboratory (VERA). Several samples showed a slightly elevated 240Pu/239Pu ratio of up to 0.22 ± 0.02 compared to global fallout (240Pu/239Pu = 0.180 ± 0.007), whereas all measured 241Pu-to-239Pu ratios were consistent with nuclear weapon fallout (241Pu/239Pu < 2.4 × 10-3), which means that no impact from the Fukushima accident was detected. From the average 241Pu-to-239Pu ratio of 8-2+3 ×10-4 at a sampling station located at a distance of 39.6 km to FDNPP, the 1-σ upper limit for the FDNPP contribution to the 239Pu inventory in the water column was estimated to be 0.2%. Pu, with the signature of weapon-grade Pu was found in a single sample collected around 770 km off the west coast of the United States.

Time-resolved 2-million-year-old supernova activity discovered in Earth's microfossil record.

Massive stars ([Formula: see text]), which terminate their evolution as core-collapse supernovae, are theoretically predicted to eject [Formula: see text] of the radioisotope (60)Fe (half-life 2.61 Ma). If such an event occurs sufficiently close to our solar system, traces of the supernova debris could be deposited on Earth. Herein, we report a time-resolved (60)Fe signal residing, at least partially, in a biogenic reservoir. Using accelerator mass spectrometry, this signal was found through the direct detection of live (60)Fe atoms contained within secondary iron oxides, among which are magnetofossils, the fossilized chains of magnetite crystals produced by magnetotactic bacteria. The magnetofossils were chemically extracted from two Pacific Ocean sediment drill cores. Our results show that the (60)Fe signal onset occurs around 2.6 Ma to 2.8 Ma, near the lower Pleistocene boundary, terminates around 1.7 Ma, and peaks at about 2.2 Ma.

Quantitative phospho-proteomics to investigate the polo-like kinase 1-dependent phospho-proteome.

Polo-like kinase 1 (PLK1) is a key regulator of mitotic progression and cell division, and small molecule inhibitors of PLK1 are undergoing clinical trials to evaluate their utility in cancer therapy. Despite this importance, current knowledge about the identity of PLK1 substrates is limited. Here we present the results of a proteome-wide analysis of PLK1-regulated phosphorylation sites in mitotic human cells. We compared phosphorylation sites in HeLa cells that were or were not treated with the PLK1-inhibitor BI 4834, by labeling peptides via methyl esterification, fractionation of peptides by strong cation exchange chromatography, and phosphopeptide enrichment via immobilized metal affinity chromatography. Analysis by quantitative mass spectrometry identified 4070 unique mitotic phosphorylation sites on 2069 proteins. Of these, 401 proteins contained one or multiple phosphorylation sites whose abundance was decreased by PLK1 inhibition. These include proteins implicated in PLK1-regulated processes such as DNA damage, mitotic spindle formation, spindle assembly checkpoint signaling, and chromosome segregation, but also numerous proteins that were not suspected to be regulated by PLK1. Analysis of amino acid sequence motifs among phosphorylation sites down-regulated under PLK1 inhibition in this data set identified two potential novel variants of the PLK1 consensus motif.

From an electrophoretic mobility shift assay to isolated transcription factors: a fast genomic-proteomic approach.

BACKGROUND: Hypocrea jecorina (anamorph Trichoderma reesei) is a filamentous ascomycete of industrial importance due to its hydrolases (e.g., xylanases and cellulases). The regulation of gene expression can influence the composition of the hydrolase cocktail, and thus, transcription factors are a major target of current research. Here, we design an approach for identifying a repressor of a xylanase-encoding gene. RESULTS: We used streptavidin affinity chromatography to isolate the Xylanase promoter-binding protein 1 (Xpp1). The optimal conditions and templates for the chromatography step were chosen according to the results of an electrophoretic mobility shift assay performed under repressing conditions, which yielded a DNA-protein complex specific to the AGAA-box (the previously identified, tetranucleotide cis-acting element). After isolating AGAA-box binding proteins, the eluted proteins were identified with Nano-HPLC/tandem MS-coupled detection. We compared the identified peptides to sequences in the H. jecorina genome and predicted in silico the function and DNA-binding ability of the identified proteins. With the results from these analyses, we eliminated all but three candidate proteins. We verified the transcription of these candidates and tested their ability to specifically bind the AGAA-box. In the end, only one candidate protein remained. We generated this protein with in vitro translation and used an EMSA to demonstrate the existence of an AGAA-box-specific protein-DNA complex. We found that the expression of this gene is elevated under repressing conditions relative to de-repressing or inducing conditions. CONCLUSIONS: We identified a putative transcription factor that is potentially involved in repressing xylanase 2 expression. We also identified two additional potential regulatory proteins that bind to the xyn2 promoter. Thus, we succeeded in identifying novel, putative transcription factors for the regulation of xylanase expression in H. jecorina.

QIKS--Quantitative identification of kinase substrates.

Signaling networks regulate cellular responses to external stimuli through post-translational modifications such as protein phosphorylation. Phosphoproteomics facilitate the large-scale identification of kinase substrates. Yet, the characterization of critical connections within these networks and the identification of respective kinases remain the major analytical challenge. To address this problem, we present a novel approach for the identification of direct kinase substrates using chemical genetics in combination with quantitative phosphoproteomics. Quantitative identification of kinase substrates (QIKS) is a novel-screening platform developed for the proteome-wide substrate-analysis of specific kinases. Here, we aimed to identify substrates of mitogen-activated protein kinase/Erk kinase (Mek1), an essential kinase in the mitogen-activated protein kinase cascade. An ATP analog-sensitive mutant of Mek1 (Mek1-as) was incubated with a cell extract from Mek1 deficient cells. Phosphorylated proteins were analyzed by LC-MS/MS of IMAC-enriched phosphopeptides, labeled differentially for relative quantification. The identification of extracellular regulated kinase 1/2 as the sole cytoplasmic substrates of MEK1 validates the applicability of this approach and suggests that QIKS could be used to identify substrates of a wide variety of kinases.

Spatially resolved positron annihilation spectroscopy on friction stir weld induced defects.

A friction stir welded (FSW) Al alloy sample was investigated by Doppler broadening spectroscopy (DBS) of the positron annihilation line. The spatially resolved defect distribution showed that the material in the joint zone becomes completely annealed during the welding process at the shoulder of the FSW tool, whereas at the tip, annealing is prevailed by the deterioration of the material due to the tool movement. This might be responsible for the increased probability of cracking in the heat affected zone of friction stir welds. Examination of a material pairing of steel S235 and the Al alloy Silafont36 by coincident Doppler broadening spectroscopy (CDBS) indicates the formation of annealed steel clusters in the Al alloy component of the sample. The clear visibility of Fe in the CDB spectra is explained by the very efficient trapping at the interface between steel cluster and bulk.

Xyr1 (xylanase regulator 1) regulates both the hydrolytic enzyme system and D-xylose metabolism in Hypocrea jecorina.

Xyr1 (xylanase regulator 1) of the ascomycete Hypocrea jecorina (anamorph Trichoderma reesei) was recently demonstrated to play an essential role in the transcriptional regulation of the xyn1 (xylanase 1-encoding) gene expression. Consequently, this study reports on the deletion of the xyr1 gene from the H. jecorina genome. Comparative studies of the growth behavior of the different mutant strains (deleted and retransformed xyr1) grown on various carbon sources pointed to the strongly reduced ability of the xyr1 deletion strain to utilize D-xylose and xylan. Transcriptional analysis of the xyl1 (D-xylose reductase 1-encoding) gene as well as measurements of corresponding enzymatic activities gave evidence that Xyr1 takes part in the control of the fungal D-xylose pathway, in particular in the regulation of D-xylose reductase. It could be demonstrated that the uptake of D-xylose into the fungal cell is uninfluenced in the Deltaxyr1 strain. Furthermore, transcriptional regulation of the major hydrolytic enzyme-encoding genes xyn1 and xyn2 (xylanases 1 and 2), cbh1 and cbh2 (cellobiohydrolases 1 and 2), and egl1 (endoglucanase 1) is strictly dependent on Xyr1. Regulation of the respective genes via Xyr1 is not affected by the substances mediating induction (xylose, xylobiose, and sophorose) and is indispensable for all modes of gene expression (basal, derepressed, and induced). Moreover, Xyr1, it was revealed, activated transcriptional regulation of inducer-providing enzymes such as beta-xylosidase BXLI and beta-glucosidase BGLI but was not shown to be involved in the regulation of BGLII.

Phosphoproteomics strategies for the functional analysis of signal transduction.

Protein phosphorylation is a key regulatory mechanism of cellular signalling processes. The analysis of phosphorylated proteins and the characterisation of phosphorylation sites under different biological conditions are some of the most challenging tasks in current proteomics research. Reduction of the sample complexity is one major step for the analysis of low-abundance kinase substrates, which can be achieved by various subcellular fractionation techniques. One strategy is the enrichment of phosphorylated proteins or peptides by immunoprecipitation or chromatography, e.g. immobilised metal affinity chromatography, prior to analysis. 2-DE gels are powerful tools for the analysis of phosphoproteins when combined with new multiplexing techniques like DIGE, phosphospecific stains, autoradiography or immunoblotting. In addition, several gel-free methods combining chromatography with highly sensitive MS have been successfully applied for the analysis of complex phosphoproteomes. Recently developed approaches like KESTREL or 'chemical genetics' and also protein microarrays offer new possibilities for the identification of specific kinase targets. This review summarises various strategies for the analyses of phosphoproteins with a special focus on the identification of novel kinase substrates.