Effects of a transient oxic period on mineralization of organic matter to CH4 and CO2 in anoxic peat incubations

Rates of organic matter mineralization in peatlands, and hence production of the greenhouse gases CH₄ and CO₂, are highly dependent on the distribution of oxygen in the peat. Using laboratory incubations of peat, we investigated the sensitivity of the anoxic production of CH₄ and CO₂ to a transient oxic period of a few weeks’ duration. Production rates during 3 successive anoxic periods were compared with rates in samples incubated in the presence of oxygen during the second period. In surface peat (5–10‐cm depth), with an initially high level of CH₄ production, oxic conditions during period 2 did not result in a lower potential CH₄ production rate during period 3, although production was delayed ～1 week. In permanently anoxic, deep peat (50–55‐cm depth) with a comparatively low initial production of CH₄, oxic conditions during period 2 resulted in zero production of CH₄ during period 3. Thus, the methanogens in surface peal—but not in deep peat—remained viable after several weeks of oxic conditions. In contrast to CH₄ production, the oxic period had a negligible effect on anoxic CO₂ production during period 3, in surface as well as deep peat. In both surface and deep peat, CO₂ production was several times higher under oxic than under anoxic conditions. However, for the first 2 weeks of oxic conditions, CO₂ production in the deep peat was very low. Still, deep peat obviously contained facultative microorganisms that, after a relatively short period, were able to maintain a considerably higher rate of organic matter mineralization under oxic than under anoxic conditions.     

Structural,Biochemical, and Computational Characterization of the Glycoside Hydrolase Family 7 Cellobiohydrolase of the Tree-killing Fungus Heterobasidion irregulare

Root rot fungi of the Heterobasidion annosum complex are the most damaging pathogens in temperate forests, and the recently sequenced Heterobasidion irregulare genome revealed over 280 carbohydrate-active enzymes. Here, H. irregulare was grown on biomass, and the most abundant protein in the culture filtrate was identified as the only family 7 glycoside hydrolase in the genome, which consists of a single catalytic domain, lacking a linker and carbohydrate-binding module. The enzyme, HirCel7A, was characterized biochemically to determine the optimal conditions for activity. HirCel7A was crystallized and the structure, refined at 1.7 Å resolution, confirms that HirCel7A is a cellobiohydrolase rather than an endoglucanase, with a cellulose-binding tunnel that is more closed than Phanerochaete chrysosporium Cel7D and more open than Hypocrea jecorina Cel7A, suggesting intermediate enzyme properties. Molecular simulations were conducted to ascertain differences in enzyme-ligand interactions, ligand solvation, and loop flexibility between the family 7 glycoside hydrolase cellobiohydrolases from H. irregulare, H. jecorina, and P. chrysosporium. The structural comparisons and simulations suggest significant differences in enzyme-ligand interactions at the tunnel entrance in the −7 to −4 binding sites and suggest that a tyrosine residue at the tunnel entrance of HirCel7A may serve as an additional ligand-binding site. Additionally, the loops over the active site in H. jecorina Cel7A are more closed than loops in the other two enzymes, which has implications for the degree of processivity, endo-initiation, and substrate dissociation. Overall, this study highlights molecular level features important to understanding this biologically and industrially important family of glycoside hydrolases.     

Parkinson Disease Protein DJ-1 Binds Metals and Protects against Metal-induced Cytotoxicity

The progressive loss of motor control due to reduction of dopamine-producing neurons in the substantia nigra pars compacta and decreased striatal dopamine levels are the classically described features of Parkinson disease (PD). Neuronal damage also progresses to other regions of the brain, and additional non-motor dysfunctions are common. Accumulation of environmental toxins, such as pesticides and metals, are suggested risk factors for the development of typical late onset PD, although genetic factors seem to be substantial in early onset cases. Mutations of DJ-1 are known to cause a form of recessive early onset Parkinson disease, highlighting an important functional role for DJ-1 in early disease prevention. This study identifies human DJ-1 as a metal-binding protein able to evidently bind copper as well as toxic mercury ions in vitro. The study further characterizes the cytoprotective function of DJ-1 and PD-mutated variants of DJ-1 with respect to induced metal cytotoxicity. The results show that expression of DJ-1 enhances the cells'' protective mechanisms against induced metal toxicity and that this protection is lost for DJ-1 PD mutations A104T and D149A. The study also shows that oxidation site-mutated DJ-1 C106A retains its ability to protect cells. We also show that concomitant addition of dopamine exposure sensitizes cells to metal-induced cytotoxicity. We also confirm that redox-active dopamine adducts enhance metal-catalyzed oxidation of intracellular proteins in vivo by use of live cell imaging of redox-sensitive S3roGFP. The study indicates that even a small genetic alteration can sensitize cells to metal-induced cell death, a finding that may revive the interest in exogenous factors in the etiology of PD.     

Catalytic Turnover Triggers Exchange of Subunits of the Magnesium Chelatase AAA+ Motor Unit

The ATP-dependent insertion of Mg²⁺ into protoporphyrin IX is the first committed step in the chlorophyll biosynthetic pathway. The reaction is catalyzed by magnesium chelatase, which consists of three gene products: BchI, BchD, and BchH. The BchI and BchD subunits belong to the family of AAA+ proteins (ATPases associated with various cellular activities) and form a two-ring complex with six BchI subunits in one layer and six BchD subunits in the other layer. This BchID complex is a two-layered trimer of dimers with the ATP binding site located at the interface between two neighboring BchI subunits. ATP hydrolysis by the BchID motor unit fuels the insertion of Mg²⁺ into the porphyrin by the BchH subunit. In the present study, we explored mutations that were originally identified in semidominant barley (Hordeum vulgare L.) mutants. The resulting recombinant BchI proteins have marginal ATPase activity and cannot contribute to magnesium chelatase activity although they apparently form structurally correct complexes with BchD. Mixing experiments with modified and wild-type BchI in various combinations showed that an exchange of BchI subunits in magnesium chelatase occurs during the catalytic cycle, which indicates that dissociation of the complex may be part of the reaction mechanism related to product release. Mixing experiments also showed that more than three functional interfaces in the BchI ring structure are required for magnesium chelatase activity.     

Dynamics and Management of Stage-Structured Fish Stocks

With increasing fishing pressures having brought several stocks to the brink of collapse, there is a need for developing efficient harvesting methods that account for factors beyond merely yield or profit. We consider the dynamics and management of a stage-structured fish stock. Our work is based on a consumer-resource model which De Roos et al. (in Theor. Popul. Biol. 73, 47–62, 2008) have derived as an approximation of a physiologically-structured counterpart. First, we rigorously prove the existence of steady states in both models, that the models share the same steady states, and that there exists at most one positive steady state. Furthermore, we carry out numerical investigations which suggest that a steady state is globally stable if it is locally stable. Second, we consider multiobjective harvesting strategies which account for yield, profit, and the recovery potential of the fish stock. The recovery potential is a measure of how quickly a fish stock can recover from a major disturbance and serves as an indication of the extinction risk associated with a harvesting strategy. Our analysis reveals that a small reduction in yield or profit allows for a disproportional increase in recovery potential. We also show that there exists a harvesting strategy with yield close to the maximum sustainable yield (MSY) and profit close to that associated with the maximum economic yield (MEY). In offering a good compromise between MSY and MEY, we believe that this harvesting strategy is preferable in most instances. Third, we consider the impact of harvesting on population size structure and analytically determine the most and least harmful harvesting strategies. We conclude that the most harmful harvesting strategy consists of harvesting both adults and juveniles, while harvesting only adults is the least harmful strategy. Finally, we find that a high percentage of juvenile biomass indicates elevated extinction risk and might therefore serve as an early-warning signal of impending stock collapse.     

PINT: a software for integration of peak volumes and extraction of relaxation rates

We present the software Peak INTegration (PINT), designed to perform integration of peaks in NMR spectra. The program is very simple to run, yet powerful enough to handle complicated spectra. Peaks are integrated by fitting predefined line shapes to experimental data and the fitting can be customized to deal with, for instance, heavily overlapped peaks. The results can be inspected visually, which facilitates systematic optimization of the line shape fitting. Finally, integrated peak volumes can be used to extract parameters such as relaxation rates and information about low populated states. The utility of PINT is demonstrated by applications to the 59 residue SH3 domain of the yeast protein Abp1p and the 289 residue kinase domain of murine EphB2.     

SSF of steam-pretreated wheat straw with the addition of saccharified or fermented wheat meal in integrated bioethanol production

Background

Integration of second-generation (2G) bioethanol production with existing first-generation (1G) production may facilitate commercial production of ethanol from cellulosic material. Since 2G hydrolysates have a low sugar concentration and 1G streams often have to be diluted prior to fermentation, mixing of streams is beneficial. Improved ethanol concentrations in the 2G production process lowers energy demand in distillation, improves overall energy efficiency and thus lower production cost. There is also a potential to reach higher ethanol yields, which is required in economically feasible ethanol production. Integrated process scenarios with addition of saccharified wheat meal (SWM) or fermented wheat meal (FWM) were investigated in simultaneous saccharification and (co-)fermentation (SSF or SSCF) of steam-pretreated wheat straw, while the possibility of recovering the valuable protein-rich fibre residue from the wheat was also studied.

Results

The addition of SWM to SSF of steam-pretreated wheat straw, using commercially used dried baker’s yeast, S. cerevisiae, resulted in ethanol concentrations of about 60 g/L, equivalent to ethanol yields of about 90% of the theoretical. The addition of FWM in batch mode SSF was toxic to baker’s yeast, due to the ethanol content of FWM, resulting in a very low yield and high accumulation of glucose. The addition of FWM in fed-batch mode still caused a slight accumulation of glucose, but the ethanol concentration was fairly high, 51.2 g/L, corresponding to an ethanol yield of 90%, based on the amount of glucose added.In batch mode of SSCF using the xylose-fermenting, genetically modified S. cerevisiae strain KE6-12, no improvement was observed in ethanol yield or concentration, compared with baker’s yeast, despite the increased xylose utilization, probably due to the considerable increase in glycerol production. A slight increase in xylose consumption was seen when glucose from SWM was fed at a low feed rate, after 48 hours, compared with batch SSCF. However, the ethanol yield and concentration remained in the same range as in batch mode.

Conclusion

Ethanol concentrations of about 6% (w/v) were obtained, which will result in a significant reduction in the cost of downstream processing, compared with SSF of the lignocellulosic substrate alone. As an additional benefit, it is also possible to recover the protein-rich residue from the SWM in the process configurations presented, providing a valuable co-product.

     

Lichenicolous fungi show population subdivision by host species but do not share population history with their hosts

Lichenicolous fungi are a species-rich biological group growing on lichen thalli. Here, we analyze the genetic structure of the lichenicolous basidiomycete Tremella lobariacearum and three host species (Lobaria pulmonaria, Lobaria macaronesica, and Lobaria immixta) in Macaronesia. We used ordination and analysis of molecular variance to investigate the structuring of genetic variation, and a simulation test to investigate whether rDNA haplotypes of T. lobariacearum were significantly associated with host species. To investigate the evolutionary and demographic history of the lichenicolous fungus and its hosts, we used coalescent samplers to generate trees, and Bayesian skyline plots. We found that the hosts were most important in structuring populations of the lichenicolous species. Despite their wide geographic distribution, the same haplotypes of T. lobariacearum consistently associated with a given host species. Our results suggest that the Lobaria hosts create a selective environment for the lichenicolous fungus. Both the pathogen and the host populations exhibited substantial genetic structure. However, evolutionary and demographic histories differed between the parasite and its hosts, as evidenced by different divergence times and tree topologies.     

Variance Heterogeneity in Saccharomyces cerevisiae Expression Data: Trans-Regulation and Epistasis

Here, we describe the results from the first variance heterogeneity Genome Wide Association Study (VGWAS) on yeast expression data. Using this forward genetics approach, we show that the genetic regulation of gene-expression in the budding yeast, Saccharomyces cerevisiae, includes mechanisms that can lead to variance heterogeneity in the expression between genotypes. Additionally, we performed a mean effect association study (GWAS). Comparing the mean and variance heterogeneity analyses, we find that the mean expression level is under genetic regulation from a larger absolute number of loci but that a higher proportion of the variance controlling loci were trans-regulated. Both mean and variance regulating loci cluster in regulatory hotspots that affect a large number of phenotypes; a single variance-controlling locus, mapping close to DIA2, was found to be involved in more than 10% of the significant associations. It has been suggested in the literature that variance-heterogeneity between the genotypes might be due to genetic interactions. We therefore screened the multi-locus genotype-phenotype maps for several traits where multiple associations were found, for indications of epistasis. Several examples of two and three locus genetic interactions were found to involve variance-controlling loci, with reports from the literature corroborating the functional connections between the loci. By using a new analytical approach to re-analyze a powerful existing dataset, we are thus able to both provide novel insights to the genetic mechanisms involved in the regulation of gene-expression in budding yeast and experimentally validate epistasis as an important mechanism underlying genetic variance-heterogeneity between genotypes.