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Targeted protein degradation systems developed for eukaryotes employ cytoplasmic machineries to perform proteolysis. This has prevented mitochondria-specific analysis of proteins that localize to multiple locations, for example, the mitochondria and the nucleus. Here, we present an inducible mitochondria-specific protein degradation system in Saccharomyces cerevisiae based on the Mesoplasma florum Lon (mf-Lon) protease and its corresponding ssrA tag (called PDT). We show that mitochondrially targeted mf-Lon protease efficiently and selectively degrades a PDT-tagged reporter protein localized to the mitochondrial matrix. The degradation can be induced by depleting adenine from the medium, and tuned by altering the promoter strength of the MF-LON gene. We furthermore demonstrate that mf-Lon specifically degrades endogenous, PDT-tagged mitochondrial proteins. Finally, we show that mf-Lon-dependent PDT degradation can also be achieved in human mitochondria. In summary, this system provides an efficient tool to selectively analyze the mitochondrial function of dually localized proteins. Conditional degradation of proteins is instrumental to advance our knowledge of cell biology but has been lacking for organelles like mitochondria. Here, the authors develop a proteolysis system based on the mycoplasma Lon protease that functions selectively within mitochondria in yeast and human cells.

Sister-chromatid cohesion, established during replication by the protein complex cohesin, is essential for both chromosome segregation and double-strand break (DSB) repair. Normally, cohesion formation is strictly limited to the S phase of the cell cycle, but DSBs can trigger cohesion also after DNA replication has been completed. The function of this damage-induced cohesion remains unknown. In this investigation, we show that damage-induced cohesion is essential for repair in postreplicative cells in yeast. Furthermore, it is established genome-wide after induction of a single DSB, and it is controlled by the DNA damage response and cohesin-regulating factors. We thus define a cohesion establishment pathway that is independent of DNA duplication and acts together with cohesion formed during replication in sister chromatid-based DSB repair.

Carboxylic acids (CAs), amino acids (AAs) and carbohydrates (CHs) in dissolved free forms can be readily assimilated by aquatic bacteria and metabolized at high growth efficiencies. Previous studies have shown that these low-molecular-weight (LMW) substrates are released by phytoplankton but also that unidentified LMW compounds of terrestrial origin is a subsidy for bacterial metabolism in unproductive freshwater systems. We tested the hypothesis that different terrestrially derived CA, AA and CH compounds can offer substantial support for aquatic bacterial metabolism in fresh waters that are dominated by allochthonous dissolved organic matter (DOM). Drainage water from three catchments of different characters in the Krycklan experimental area in Northern Sweden were studied at the rising and falling limb of the spring flood, using a 2-week bioassay approach. A variety of CA, AA and CH compounds were significantly assimilated by bacteria, meeting 15–100% of the bacterial carbon demand and explaining most of the observed variation in bacterial growth efficiency (BGE; R 2 =0.66). Of the 29 chemical species that was detected, acetate was the most important, representing 45% of the total bacterial consumption of all LMW compounds. We suggest that LMW organic compounds in boreal spring flood drainage could potentially support all in situ bacterial production in receiving lake waters during periods of weeks to months after the spring flood.

Permafrost methane: Arctic emissions revisited A late-autumn 'shoulder' is a regular feature of the seasonal cycles of atmospheric methane at high latitudes, but the sources of this burst of methane remain obscure. Mastepanov et al. now report methane flux measurements from a high Arctic setting during the onset of soil freezing. The total emissions during this freeze-in period are roughly equal to the amount of methane emitted during the entire summer season. Including the observed methane burst in an atmospheric chemistry and transport model improves agreement between the simulated seasonal cycle and atmospheric data from latitudes north of 600N. These results suggest that permafrost-associated freeze-in bursts of methane from tundra regions may be an important, previously unrecognized component of the seasonal distribution of methane emissions at high latitudes. A late-autumn shoulder is consistently observed in the seasonal cycles of atmospheric methane at high latitude sites, but the sources responsible remain uncertain. This study reports methane flux measurements from a high Arctic setting during the onset of soil freezing. The integral of the emissions during this freeze-in period amount to approximately the same amount of methane emitted during the entire summer season. It is found that the observed early winter emission burst improves the agreement between the simulated seasonal cycle and atmospheric data from latitudes north of 60°N. The results suggest that permafrost associated freeze-in bursts of methane emissions from tundra regions could be an important component of the seasonal distribution of methane emissions from high latitudes. Terrestrial wetland emissions are the largest single source of the greenhouse gas methane 1 . Northern high-latitude wetlands contribute significantly to the overall methane emissions from wetlands, but the relative source distribution between tropical and high-latitude wetlands remains uncertain 2 , 3 . As a result, not all the observed spatial and seasonal patterns of atmospheric methane concentrations can be satisfactorily explained, particularly for high northern latitudes. For example, a late-autumn shoulder is consistently observed in the seasonal cycles of atmospheric methane at high-latitude sites 4 , but the sources responsible for these increased methane concentrations remain uncertain. Here we report a data set that extends hourly methane flux measurements from a high Arctic setting into the late autumn and early winter, during the onset of soil freezing. We find that emissions fall to a low steady level after the growing season but then increase significantly during the freeze-in period. The integral of emissions during the freeze-in period is approximately equal to the amount of methane emitted during the entire summer season. Three-dimensional atmospheric chemistry and transport model simulations of global atmospheric methane concentrations indicate that the observed early winter emission burst improves the agreement between the simulated seasonal cycle and atmospheric data from latitudes north of 60° N. Our findings suggest that permafrost-associated freeze-in bursts of methane emissions from tundra regions could be an important and so far unrecognized component of the seasonal distribution of methane emissions from high latitudes.

Abstract Background Occurrence of low blood taurine concentrations (B-TauC) and predisposing factors to taurine deficiency in English Cocker Spaniels (ECS) are incompletely understood. Objectives Investigate the occurrence of low B-TauC in a Swedish population of ECS and evaluate the association between B-TauC and dog characteristics, clinical variables, and diet composition. Animals One-hundred eighty privately owned ECS. Methods Dogs were prospectively recruited and underwent physical examination, blood analyses, and echocardiographic and ophthalmic examinations. Dogs with clinical signs of congestive heart failure (CHF) also underwent thoracic radiography. Taurine concentrations were analyzed in plasma (EDTA and heparin) and whole blood. Diets consumed by the dogs at the time of the examination were analyzed for dietary taurine- (D-TauC), cysteine- (D-CysC), and methionine concentrations (D-MetC). Results Fifty-three of 180 dogs (29%) had low B-TauC, of which 13 (25%) dogs had clinical and radiographic signs of CHF, increased echocardiographic left ventricular (LV) dimensions and volumes, and impaired LV systolic function. Five (9%) dogs with low B-TauC had retinal abnormalities. Dietary MetC, dietary animal protein source (red/white meat), and age were associated with B-TauC in the final multivariable regression model (P < .001, R2adj = .39). Conclusions and Clinical Importance Low B-TauC suggests that taurine deficiency may play a role in the development of myocardial failure and CHF in ECS. Low D-MetC and diets with red meat as the animal protein source were associated with low B-TauC. Dogs with B-TauC below the normal reference range were older than dogs with normal concentrations.

Canine ovariectomy is a commonly performed procedure in veterinary practice and can be done via an open celiotomy or a laparoscopic approach. Laparoscopy requires different psychomotor skills from those involved in open surgery. Many of these basic skills can initially be acquired through simulation-based training. Surgical simulation training benefits both the surgeon and the patient by offering the acquisition of new skills in a less stressful environment while lowering the risk for the patient. Virtual reality (VR) simulation training is well established in human medicine, but no VR modules have previously been available for veterinary procedure training for laparoscopy. This project aimed to develop a VR simulator module for canine laparoscopic ovariectomy on the simulator LapSim ® (Surgical Science, Gothenburg, Sweden). The simulator module could offer veterinary surgeons the opportunity to gain the basic technical skills required for canine laparoscopic ovariectomy. Surgeons with experience in laparoscopic ovariectomy acted as content experts, and the development was a stepwise iterative process. The three-port technique was chosen; it included locating and lifting the uterine horn and sealing and transecting tissue to free the ovaries. The organs and tissues the surgeon interacted with during the simulation were the spleen, intestines, pancreas, ovaries, uterine horns, suspensory ligaments, mesometrium and mesovarium. The module included simulating haemorrhage from the spleen, ovarian and uterine vessels. Suggestions for the continued development of the simulation were identified. Examples were an increased haptic feel in some instances of tissue manipulation and the possibility of introducing a variation in the length of the ovarian pedicle. The last step of the simulated procedure included visual guidance for verification of haemostasis. Even though guidance is not part of a real-life operation, identifying anatomical landmarks was considered valuable from an educational perspective. A validity investigation study has been initiated to evaluate the module as an assessment tool.

Herbivory is an important part of most ecosystems and affects the ecosystems' carbon balance both directly and indirectly. Little is known about herbivory and its impact on the carbon balance in high arctic mire ecosystems. We hypothesized that trampling and grazing by large herbivores influences the vegetation density and composition and thereby also the carbon balance. In 2010, we established fenced exclosures in high arctic Greenland to prevent muskoxen (Ovibos moschatus) from grazing. During the growing seasons of 2011 to 2013 we measured CO2 and CH4 fluxes in these ungrazed blocks and compared them to blocks subjected to natural grazing. Additionally, we measured depth of the water table and active layer, soil temperature, and in 2011 and 2013 an inventory of the vegetation density and composition were made. In 2013 a significant decrease in total number of vascular plant (33-44%) and Eriophorum scheuchzeri (51-53%) tillers were found in ungrazed plots, the moss-layer and amount of litter had also increased substantially in these plots. This resulted in a significant decrease in net ecosystem uptake of CO2 (47%) and likewise a decrease in CH4 emission (44%) in ungrazed plots in 2013. While the future of the muskoxen in a changing arctic is unknown, this experiment points to a potentially large effect of large herbivores on the carbon balance in natural Arctic ecosystems. It thus sheds light on the importance of grazing mammals, and hence adds to our understanding of natural ecosystem greenhouse gas balance in the past and in the future.

A liquid chromatography/time-of-flight mass spectrometry (LC-TOF-MS) method for targeted toxicological screening in human postmortem blood samples from forensic autopsy cases has been developed, validated and compared with a previously used method using gas chromatography with nitrogen–phosphorus detection (GC-NPD). Separation was achieved within 12 min by high-resolution gradient chromatography. Ions were generated in positive and negative electrospray ionization mode and were detected in 2-GHz single mass spectrometry mode, m / z range 50–1,000. Before injection, 0.25 g blood was prepared by protein precipitation with 500 μL of a mixture of acetonitrile and ethanol containing deuterated internal standards. An in-house database comprising 240 drugs and metabolites was built by analysing solutions from certified standards or other documented reference material available. Identification was based on scoring of retention time, accurate mass measurement and isotopic pattern. Validation was performed on spiked blood samples and authentic postmortem blood samples. The thresholds defined as minimum required performance levels were for most compounds in the range from 0.01 to 0.10 μg/g. Typically, a mass error of less than 2 ppm and a precision of area measurements of less than 5 % coefficient of variation were achieved. Positive identification was confirmed at concentrations up to 500 μg/g. Most compounds were determined in positive ionization mode, but for a limited number of compounds (fewer than 4 %) negative ionization was needed and a few early-eluted compounds could not be identified owing to substantial influence of interferences from the matrix and were thus not included in the screening. A robust and valid toxicological screening by LC-TOF-MS for postmortem blood samples, covering 50 % more compounds, and with higher precision and sensitivity than the previously used screening by GC-NPD was achieved.

In this study, the authors use measures of carbon-14 in neuronal DNA from human stroke patient cortical tissue samples to show that, unlike previous studies done in rodents, they do not find any evidence of increased neurogenesis after an ischemic injury. In addition, DNA damage assays suggest that there is no increase in DNA rearrangement after this insult. It has been unclear whether ischemic stroke induces neurogenesis or neuronal DNA rearrangements in the human neocortex. Using immunohistochemistry; transcriptome, genome and ploidy analyses; and determination of nuclear bomb test–derived 14 C concentration in neuronal DNA, we found neither to be the case. A large proportion of cortical neurons displayed DNA fragmentation and DNA repair a short time after stroke, whereas neurons at chronic stages after stroke showed DNA integrity, demonstrating the relevance of an intact genome for survival.

Although much attention in recent years has been devoted to methane (CH4) emissions from northern wetlands, measurement based data sets providing full annual budgets are still limited in number. This study was designed to help fill the gap of year‐round measurements of CH4 emissions from subarctic mires. We report continuous eddy correlation CH4 flux measurements made during 2006 and 2007 over the Stordalen mire in subarctic Sweden (68°20′N, 19°03′E, altitude 351 m) using a cryocooled tunable diode laser. The landscape‐scale CH4 fluxes originated mainly from the permafrost free wet parts of the mire dominated by tall graminoid vegetation. The midseason average CH4 emission mean was 6.2 ± 2.6 mg m−2 h−1. A detailed footprint analysis indicates an additional strong influence on the flux by the nearby shallow Lake Villasjön (0.17 km2, maximum depth 1.3 m). A stable bimodal distribution of wind flow from either the east or the west allowed separating the lake and mire vegetation signals. The midseason lake emission rates were as high as 12.3 ± 3.3 mg m−2 h−1. Documented CH4 fluxes are similar to results obtained by automatic chamber technique and higher than manual chamber measurements made in the wet minerotrophic section dominated by Eriophorum angustifolium. The high fluxes observed from this vegetation type are significant because the areal distribution of this source in the mire is expanding due to ongoing thawing of the permafrost. A simple peat temperature relationship with CH4 emissions was used to fill data gaps to construct a complete annual budget of CH4 fluxes over the studied area. The calculated annual CH4 emissions in 2006 and 2007 equaled 24.5 and 29.5 g CH4 m−2 yr−1, respectively. The summer season CH4 emissions dominated (65%) the annual flux, with the shoulder seasons of spring and autumn significant (25%) and a minor flux from the winter (10%).