Secretion of the Hormoconis resinae glucoamylase P enzyme from Trichoderma reesei directed by the natural and the cbh1 gene secretion signal

Abstract We have isolated two alkaline phosphatases (H-AP and L-AP, for high and low molecular mass, respectively) from Pseudomonas aeruginosa PA01. These two enzymes were found to differ in mobility on sodium dodecyl sulphate polyacrylamide gels (H-AP, M _r = 51 000 and L-AP, M _r = 39 500), amino-terminal amino acid sequence and did not cross-react. Both enzymes were active as phosphomonoesterases while only L-AP demonstrated any phosphodiesterase activity. Both enzymes were purified from P. aeruginosa grown in phosphate limiting conditions using the same protocol and were identified in both periplasmic and extracellular locations. A low level of H-AP was produced constitutively whereas L-AP was produced only after induction by reduced phosphate concentration in the growth medium. An L-AP-like enzyme has been previously described, however, this is the first report of a second P. aeruginosa alkaline phosphatase.     

Microhabitat use and behavior of voles under weasel and raptor predation risk: predator facilitation?

An example of predator facilitation is that a microhabitat shiftin a prey species induced by one predator increases the probabilityof the prey falling victim to other predators. Least weasels(Mustela nivalis) hunt in dense plant cover, whereas kestrels(Falco tinnunculus) hunt in habitats with sparse plant cover.Field voles (Microtus agrestis), the main food of weasels andkestrels, prefer open country with a high grass layer. We simulateda multipredator environment in an aviary (3.0 x 4.8 x 2.2 m)to find out whether predator facilitation plays a role in theinteractions between voles, small mustelids, and raptors. Ineach replicate, we placed a field vole in a pen including sidesof high and low grass layers (cover and open). In a predator-freesituation, voles preferred cover but shifted to open when aweasel was introduced to cover. In the presence of a kestrel,voles occupied cover and decreased their mobility. In the presenceof a weasel plus a kestrel, voles behaved as under the kestrelrisk alone. Therefore, in these aviary circumstances, volesperceived the kestrel risk as greater than the weasel risk.Predator facilitation in the assemblage of predators subsistingon rodent prey may contribute to the crash of the four-yearvole cycle: microhabitat shift due to an avoidance of weaseljaws may drive voles to raptor talons.     

Eco-efficiency indicator framework implemented in the metallurgical industry: part 1—a comprehensive view and benchmark

Purpose

The purpose of this work was to develop an indicator framework for the environmental sustainability benchmarking of products produced by the metallurgical industry. Sustainability differentiation has become an important issue for companies throughout the value chain. Differentiation is sometimes not attainable, due to the use of average data, lack of comparative data, certain issues being overshadowed by others, and a very narrow palette of indicators dominating the current sustainability assessments. There is a need for detailed and credible analyses, which show the current status and point out where improvements can be made. The indicator framework is developed to give a comprehensive picture of eco-efficiency, to provide methods that enable relevant comparisons as well as the tools for communicating the results. In this way, the methodology presented in this study aims to make differentiation easier and thus aid companies in driving the development toward more sustainable solutions.

Methods

The framework is based on the existing indicator framework Gaia Biorefiner, which is primarily intended for bio-based products. In this work, the framework was further developed for application in the metallurgical industry. The indicator framework is built by first looking at the issues, which are critical to the environment and global challenges seen today and which the activities of the metallurgical industry may have an impact on. Based on these issues, suitable indicators are chosen if they exist and built if they do not. The idea is that all indicators in a group form a whole, showing areas of innovation while refraining from aggregating and weighting, which often compromise a comprehensive and objective view. Both qualitative and quantitative indicators are included. The indicators are constructed following the criteria set by the EU and OECD for building indicators. Each indicator further has a benchmark. The rules for building the benchmark are connected to the indicators. Suitable data sources and criteria for the benchmark and the indicators are gathered from literature, publicly available databases, and commercial LCA software. The use of simulation tools for attaining more reliable data is also studied.

Results and discussion

The result is a visual framework consisting of ten indicator groups with one to five indicators each, totaling up to 31 indicators. These are visualized in a sustainability indicator “flower.” The flower can be further opened up to study each indicator and the reasons behind the results. The sustainability benchmark follows a methodology that is based on utilization of baseline data and sustainability criteria or limits. A simulation approach was included in the methodology to address the problem with data scarcity and data reliability. The status of the environment, current production technologies, location-specific issues, and process-specific issues all affect the result, and the aim of finding relevant comparisons that will support sustainability differentiation is answered by a scalable scoping system.

Conclusions

A new framework and its concise visualization has been built for assessing the eco-efficiency of products from the metallurgical industry, in a way that aims to answer the needs of the industry. Since there is a baseline, against which each indicator can be benchmarked, a sustainability indicator “flower” can be derived, one of the key innovations of this methodology. This approach goes beyond the usual quantification, as it is also scalable and linked to technology and its fundamental parameters. In part 2, a case study “A case study from the copper industry” tests and illustrates the methodology.

     

Circulation pattern and transmission dynamics of the supra-population of the nematode Cystidicoloides tenuissima (Zeder) in the River Swincombe, England

The circulation pattern and transmission dynamics of larval and adult Cystidicoloides tenuissima in all its intermediate and definitive hosts were investigated and quantified at three sites in a small upland stream over a period of 1 year. Both brown trout, Salmo trutta , and salmon parr, S. salar , were suitable definitive hosts, but because of the greater importance of the mayfly intermediate host in the diet of trout, between 73 and 98% of the parasite infrapopulation in fish circulated through this species. Trout was the only required host, and was alone responsible for the perpetuation of the parasite suprapopulation in the river. Around 99% of the parasite's eggs produced originated from trout but, of these, 90% or more failed to be ingested by an insect. Larval parasites were found in 18 species of insects, but could develop to the infective third stage in only one species, the mayfly Leptophlebia marginata. Two common but unsuitable species harboured up to 80% of the larval parasites, and less than 10% actually circulated through L. marginata. Differences in circulation pattern between sites could be related more to differences in fish feeding preferences than to differences in fish or insect density, but monthly differences in transmission rate reflected both fish diet and insect abundance. The overall mean transmission rate of eggs to larvae in L. marginata varied between 0.25 and 0.87%, but transmission rates of these larvae to fish were far higher, from 10.8 to 39.8%. The relative importance of ecological factors, host community structure and parasite specificity in determining circulation routes and transmission efficiencies are discussed.     

Female-biased natal dispersal in the Siberian flying squirrel

Natal dispersal is usually sex biased in birds and mammals.Female-biased natal dispersal is the prevailing pattern in birdsbut is rare among mammals. Hypotheses explaining sex bias indispersal include the mate-defense mating hypothesis, whichpredicts male-biased dispersal, the resource-defense hypothesispredicting female-biased dispersal, and the competition hypothesis,which predicts that if dispersal is caused by competition forresources between sexes, then the subdominant sex will disperse.We studied natal dispersal of Siberian flying squirrels Pteromysvolans using radio telemetry in Southern Finland in 1996–2004.Of 86 juveniles that survived over the dispersal period, almostall young females dispersed from the natal site, whereas almost40% of males were philopatric. Dispersal was farther for femalesthan males. Females began dispersal on average 2 weeks earlierthan males and were lighter in mass at the onset of dispersalthan later dispersing males. No mate- or resource-defense matingsystem could be found among males, but females seemed to defendnest and apparently food resources, in contrast to the expectationof dispersal bias in resource-defense systems. Competition forresources between sexes does not explain female bias either:in the flying squirrel, the female seems to be the dominantsex. We propose that young females are subordinate to theirmothers and have to disperse to find a vacant, suitable sitefor reproduction.     

Type II Transmembrane Serine Protease Gene Variants Associate with Breast Cancer

Type II transmembrane serine proteases (TTSPs) are related to tumor growth, invasion, and metastasis in cancer. Genetic variants in these genes may alter their function, leading to cancer onset and progression, and affect patient outcome. Here, 464 breast cancer cases and 370 controls were genotyped for 82 single-nucleotide polymorphisms covering eight genes. Association of the genotypes was estimated against breast cancer risk, breast cancer–specific survival, and survival in different treatment groups, and clinicopathological variables. SNPs in TMPRSS3 (rs3814903 and rs11203200), TMPRSS7 (rs1844925), and HGF (rs5745752) associated significantly with breast cancer risk (P _trend = 0.008–0.042). SNPs in TMPRSS1 (rs12151195 and rs12461158), TMPRSS2 (rs2276205), TMPRSS3 (rs3814903), and TMPRSS7 (rs2399403) associated with prognosis (P = 0.004–0.046). When estimating the combined effect of the variants, the risk of breast cancer was higher with 4–5 alleles present compared to 0–2 alleles (P = 0.0001; OR, 2.34; 95% CI, 1.39–3.94). Women with 6–8 survival-associating alleles had a 3.3 times higher risk of dying of breast cancer compared to women with 1–3 alleles (P = 0.001; HR, 3.30; 95% CI, 1.58–6.88). The results demonstrate the combined effect of variants in TTSPs and their related genes in breast cancer risk and patient outcome. Functional analysis of these variants will lead to further understanding of this gene family, which may improve individualized risk estimation and development of new strategies for treatment of breast cancer.     

Glucoamylase P gene of Hormoconis resinae: Molecular cloning, sequencing and introduction into Trichoderma reesei

The glucoamylase P gene of the fungus Hormoconis resinae has been cloned and sequenced from a genomic library. The gene consists of a 2153-bp protein coding region including three introns. The usual number of introns in cloned fungal glucoamylase genes has been four and in some cases five. Two of the glucoamylase P gene introns contain a sequence resembling the consensus sequence found near the 3' splice site in the introns of the fungus Trichoderma reesei cellobiohydrolase 1 (cbh1) gene. The H. resinae glucoamylase P gene, under its own promoter, was introduced into T. reesei, but no expression could be detected.     

DNA family shuffling within the chicken avidin protein family - A shortcut to more powerful protein tools

Avidins represent an interesting group of proteins showing high structural similarity and ligand-binding properties but low similarity in primary structure. In this study, we show that it is possible to create functional chimeric proteins from the avidin protein family when applying DNA family shuffling to the genes of the avidin protein family: avidin, avidin related gene 2 and biotin-binding protein A. The novel chimeric proteins were selected by phage display biopanning against biotin, and the selected enriched proteins were characterized, displaying diverse features distinct from the parental genes, including binding to cysteine.     

Epithelial and ectomesenchymal role of the type I TGF-beta receptor ALK5 during facial morphogenesis and palatal fusion

Transforming growth factor beta (TGF-beta) proteins play important roles in morphogenesis of many craniofacial tissues; however, detailed biological mechanisms of TGF-beta action, particularly in vivo, are still poorly understood. Here, we deleted the TGF-beta type I receptor gene Alk5 specifically in the embryonic ectodermal and neural crest cell lineages. Failure in signaling via this receptor, either in the epithelium or in the mesenchyme, caused severe craniofacial defects including cleft palate. Moreover, the facial phenotypes of neural crest-specific Alk5 mutants included devastating facial cleft and appeared significantly more severe than the defects seen in corresponding mutants lacking the TGF-beta type II receptor (TGFbetaRII), a prototypical binding partner of ALK5. Our data indicate that ALK5 plays unique, non-redundant cell-autonomous roles during facial development. Remarkable divergence between Tgfbr2 and Alk5 phenotypes, together with our biochemical in vitro data, imply that (1) ALK5 mediates signaling of a diverse set of ligands not limited to the three isoforms of TGF-beta, and (2) ALK5 acts also in conjunction with type II receptors other than TGFbetaRII.     

ALS-linked P56S-VAPB, an aggregated loss-of-function mutant of VAPB, predisposes motor neurons to ER stress-related death by inducing aggregation of co-expressed wild-type VAPB

A point mutation (P56S) in the vapb gene encoding an endoplasmic reticulum (ER)-integrated membrane protein [vesicle-associated membrane protein-associated protein B (VAPB)] causes autosomal-dominant amyotrophic lateral sclerosis. In our earlier study, we showed that VAPB may be involved in the IRE1/XBP1 signaling of the unfolded protein response, an ER reaction to inhibit accumulation of unfolded/misfolded proteins, while P56S-VAPB formed insoluble aggregates and lost the ability to mediate the pathway (loss-of-function), and suggested that P56S-VAPB promoted the aggregation of co-expressed wild-type (wt)-VAPB. In this study, a yeast inositol-auxotrophy assay has confirmed that P56S-VAPB is functionally a null mutant in vivo . The interaction between P56S-VAPB and wt-VAPB takes place with a high affinity through the major sperm protein domain in addition to the interaction through the C-terminal transmembrane domain. Consequently, wt-VAPB is speculated to preferentially interact with co-expressed P56S-VAPB, leading to the recruitment of wt-VAPB into cytosolic aggregates and the attenuation of its normal function. We have also found that expression of P56S-VAPB increases the vulnerability of NSC34 motoneuronal cells to ER stress-induced death. These results lead us to hypothesize that the total loss of VAPB function in unfolded protein response, induced by one P56S mutant allele, may contribute to the development of P56S-VAPB-induced amyotrophic lateral sclerosis.