Secretion and assembly of functional mini-cellulosomes from synthetic chromosomal operons in Clostridium acetobutylicum ATCC 824.

Background: Consolidated bioprocessing (CBP) is reliant on the simultaneous enzyme production, saccharification of biomass, and fermentation of released sugars into valuable products such as butanol. Clostridial species that produce butanol are, however, unable to grow on crystalline cellulose. In contrast, those saccharolytic species that produce predominantly ethanol, such as Clostridium thermocellum and Clostridium cellulolyticum, degrade crystalline cellulose with high efficiency due to their possession of a multienzyme complex termed the cellulosome. This has led to studies directed at endowing butanol-producing species with the genetic potential to produce a cellulosome, albeit by localising the necessary transgenes to unstable autonomous plasmids. Here we have explored the potential of our previously described Allele-Coupled Exchange (ACE) technology for creating strains of the butanol producing species Clostridium acetobutylicum in which the genes encoding the various cellulosome components are stably integrated into the genome. Results: We used BioBrick2 (BB2) standardised parts to assemble a range of synthetic genes encoding C. thermocellum cellulosomal scaffoldin proteins (CipA variants) and glycoside hydrolases (GHs, Cel8A, Cel9B, Cel48S and Cel9K) as well as synthetic cellulosomal operons that direct the synthesis of Cel8A, Cel9B and a truncated form of CipA. All synthetic genes and operons were integrated into the C. acetobutylicum genome using the recently developed ACE technology. Heterologous protein expression levels and mini-cellulosome self-assembly were assayed by western blot and native PAGE analysis. Conclusions: We demonstrate the successful expression, secretion and self-assembly of cellulosomal subunits by the recombinant C. acetobutylicum strains, providing a platform for the construction of novel cellulosomes. © 2013 Kovács et al.; licensee BioMed Central Ltd

Towards the reconstruction of integrated genome-scale models of metabolism and gene expression

The reconstruction of integrated genome-scale models of metabolism and gene expression has been a challenge for a while now. In fact, various methods that allow integrating reconstructions of Transcriptional Regulatory Networks, gene expression data or both into Genome-Scale Metabolic Models have been proposed. Several of these methods are surveyed in this article, which allowed identifying their strengths and weaknesses concerning the reconstruction of integrated models for multiple prokaryotic organisms. Additionally, the main resources of regulatory information were also surveyed, as the existence of novel sources of regulatory information and gene expression data may contribute for the improvement of methodologies referred herein.This study was supported by the Portuguese Foundation for Science andTechnology (FCT) under the scope of the strategic funding of UID/BIO/04469/2019 unit andBioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by the European RegionalDevelopment Fund under the scope of Norte2020-Programa Operacional Regional do Norte. Fernando Cruz holds a doctoral fellowship (SFRH/BD/139198/2018) funded by the FCT. The authors thank project SHIKIFACTORY100 - Modular cell factories for the production of 100 compounds from the shikimate pathway (814408) funded by the European Commission.info:eu-repo/semantics/publishedVersio

Clusters of galaxies : observational properties of the diffuse radio emission

Clusters of galaxies, as the largest virialized systems in the Universe, are ideal laboratories to study the formation and evolution of cosmic structures...(abridged)... Most of the detailed knowledge of galaxy clusters has been obtained in recent years from the study of ICM through X-ray Astronomy. At the same time, radio observations have proved that the ICM is mixed with non-thermal components, i.e. highly relativistic particles and large-scale magnetic fields, detected through their synchrotron emission. The knowledge of the properties of these non-thermal ICM components has increased significantly, owing to sensitive radio images and to the development of theoretical models. Diffuse synchrotron radio emission in the central and peripheral cluster regions has been found in many clusters. Moreover large-scale magnetic fields appear to be present in all galaxy clusters, as derived from Rotation Measure (RM) studies. Non-thermal components are linked to the cluster X-ray properties, and to the cluster evolutionary stage, and are crucial for a comprehensive physical description of the intracluster medium. They play an important role in the cluster formation and evolution. We review here the observational properties of diffuse non-thermal sources detected in galaxy clusters: halos, relics and mini-halos. We discuss their classification and properties. We report published results up to date and obtain and discuss statistical properties. We present the properties of large-scale magnetic fields in clusters and in even larger structures: filaments connecting galaxy clusters. We summarize the current models of the origin of these cluster components, and outline the improvements that are expected in this area from future developments thanks to the new generation of radio telescopes.Comment: Accepted for the publication in The Astronomy and Astrophysics Review. 58 pages, 26 figure

A Retrospective Cohort Study of the Potency of lipid-lowering therapy and Race-gender Differences in LDL cholesterol control

<p>Abstract</p> <p>Background</p> <p>Reasons for race and gender differences in controlling elevated low density lipoprotein (LDL) cholesterol may be related to variations in prescribed lipid-lowering therapy. We examined the effect of lipid-lowering drug treatment and potency on time until LDL control for black and white women and men with a baseline elevated LDL.</p> <p>Methods</p> <p>We studied 3,484 older hypertensive patients with dyslipidemia in 6 primary care practices over a 4-year timeframe. Potency of lipid-lowering drugs calculated for each treated day and summed to assess total potency for at least 6 and up to 24 months. Cox models of time to LDL control within two years and logistic regression models of control within 6 months by race-gender adjust for: demographics, clinical, health care delivery, primary/specialty care, LDL measurement, and drug potency.</p> <p>Results</p> <p>Time to LDL control decreased as lipid-lowering drug potency increased (P < 0.001). Black women (N = 1,440) received the highest potency therapy (P < 0.001) yet were less likely to achieve LDL control than white men (N = 717) (fully adjusted hazard ratio [HR] 0.66 [95% CI 0.56-0.78]). Black men (N = 666) and white women (N = 661) also had lower adjusted HRs of LDL control (0.82 [95% CI 0.69, 0.98] and 0.75 [95% CI 0.64-0.88], respectively) than white men. Logistic regression models of LDL control by 6 months and other sensitivity models affirmed these results.</p> <p>Conclusions</p> <p>Black women and, to a lesser extent, black men and white women were less likely to achieve LDL control than white men after accounting for lipid-lowering drug potency as well as diverse patient and provider factors. Future work should focus on the contributions of medication adherence and response to treatment to these clinically important differences.</p

New Strategy for Rapid Diagnosis and Characterization of Fungal Infections: The Example of Corneal Scrapings

PURPOSE: The prognosis of people infected with Fungi especially immunocompromised depends on rapid and accurate diagnosis to capitalize on time administration of specific treatments. However, cultures produce false negative results and nucleic-acid amplification techniques require complex post-amplification procedures to differentiate relevant fungal types. The objective of this work was to develop a new diagnostic strategy based on real-time polymerase-chain reaction high-resolution melting analysis (PCR-HRM) that a) detects yeasts and filamentous Fungi, b) differentiates yeasts from filamentous Fungi, and c) discriminates among relevant species of yeasts. METHODS: PCR-HRM detection limits and specificity were assessed with a) isolated strains; b) human blood samples experimentally infected with Fungi; c) blood experimentally infected with other infectious agents; d) corneal scrapings from patients with suspected fungal keratitis (culture positive and negative) and e) scrapings from patients with suspected bacterial, viral or Acanthamoeba infections. The DNAs were extracted and mixed with primers diluted in the MeltDoctor® HRM Master Mix in 2 tubes, the first for yeasts, containing the forward primer CandUn (5'CATGCCTGTTTGAGCGTC) and the reverse primer FungUn (5'TCCTCCGCTT ATTGATATGCT) and the second for filamentous Fungi, containing the forward primer FilamUn (5'TGCCTGTCCGAGCGTCAT) and FungUn. Molecular probes were not necessary. The yields of DNA extraction and the PCR inhibitors were systematically monitored. RESULTS: PCR-HRM detected 0.1 Colony Forming Units (CFU)/µl of yeasts and filamentous Fungi, differentiated filamentous Fungi from yeasts and discriminated among relevant species of yeasts. PCR-HRM performances were higher than haemoculture and sensitivity and specificity was 100% for culture positive samples, detecting and characterizing Fungi in 7 out 10 culture negative suspected fungal keratitis. CONCLUSIONS: PCR-HRM appears as a new, sensitive, specific and inexpensive test that detects Fungi and differentiates filamentous Fungi from yeasts. It allows direct fungal detection from clinical samples and experimentally infected blood in less than 2.30 h after DNA extraction

Lack of efficacy of troglitazone at clinically achievable concentrations, with or without 9-cis retinoic acid or cytotoxic agents, for hepatocellular carcinoma cell lines

[[abstract]]Although the PPARgamma agonist troglitazone has been shown to induce growth inhibition of hepatocellular carcinoma (HCC) cells at high concentration, this study indicates troglitazone does not significantly inhibit the growth of HCC cells at clinically achievable concentrations (1-10 muM), and this lack of activity could not be improved by the addition of 9-cis-retinoic acid. Furthermore, no synergistic effect was found between troglitazone and cytotoxic anticancer agents

Does Landscape Fragmentation Influence Sex Ratio of Dioecious Plants? A Case Study of Pistacia chinensis in the Thousand-Island Lake Region of China

The Thousand-Island Lake region in Zhejiang Province, China is a highly fragmented landscape with a clear point-in-time of fragmentation as a result of flooding to form the reservoir. Islands in the artificial lake were surveyed to examine how population sex ratio of a dioecious plant specie Pistacia chinensis B. was affected by landscape fragmentation. A natural population on the mainland near the lake was also surveyed for comparison. Population size, sex ratio and diameter at breast height (DBH) of individuals were measured over 2 years. More than 1,500 individuals, distributed in 31 populations, were studied. Soil nitrogen in the different populations was measured to identify the relationship between sex ratio and micro-environmental conditions. In accordance with the results of many other reports on biased sex ratio in relation to environmental gradient, we found that poor soil nitrogen areas fostered male-biased populations. In addition, the degree of sex ratio bias increased with decreasing population size and population connectivity. The biased sex ratios were only found in younger individuals (less than 50 years old) in small populations, while a stable 1∶1 sex ratio was found in the large population on the mainland. We concluded that the effects of landscape fragmentation on the dioecious population sex ratio were mainly achieved in relation to changing soil nitrogen conditions in patches and pollen limitation within and among populations. Large populations could maintain a more suitable environment in terms of nutrient conditions and pollen flow, subsequently maintaining a stable sex ratio in dioecious plant populations. Both micro-environmental factors and spatial structure should be considered in fragmented landscape for the conservation of dioecious plant species

The ε3 and ε4 Alleles of Human APOE Differentially Affect Tau Phosphorylation in Hyperinsulinemic and Pioglitazone Treated Mice

Impaired insulin signalling is increasingly thought to contribute to Alzheimer's disease (AD). The ε4 isoform of the APOE gene is the greatest genetic risk factor for sporadic, late onset AD, and is also associated with risk for type 2 diabetes mellitus (T2DM). Neuropathological studies reported the highest number of AD lesions in brain tissue of ε4 diabetic patients. However other studies assessing AD pathology amongst the diabetic population have produced conflicting reports and have failed to show an increase in AD-related pathology in diabetic brain. The thiazolidinediones (TZDs), peroxisome proliferator-activated receptor gamma agonists, are peripheral insulin sensitisers used to treat T2DM. The TZD, pioglitazone, improved memory and cognitive functions in mild to moderate AD patients. Since it is not yet clear how apoE isoforms influence the development of T2DM and its progression to AD, we investigated amyloid beta and tau pathology in APOE knockout mice, carrying human APOEε3 or ε4 transgenes after diet-induced insulin resistance with and without pioglitazone treatment.Male APOE knockout, APOEε3-transgenic and APOEε4-transgenic mice, together with background strain C57BL6 mice were kept on a high fat diet (HFD) or low fat diet (LFD) for 32 weeks, or were all fed HFD for 32 weeks and during the final 3 weeks animals were treated with pioglitazone or vehicle.All HFD animals developed hyperglycaemia with elevated plasma insulin. Tau phosphorylation was reduced at 3 epitopes (Ser396, Ser202/Thr205 and Thr231) in all HFD, compared to LFD, animals independent of APOE genotype. The introduction of pioglitazone to HFD animals led to a significant reduction in tau phosphorylation at the Ser202/Thr205 epitope in APOEε3 animals only. We found no changes in APP processing however the levels of soluble amyloid beta 40 was reduced in APOE knockout animals treated with pioglitazone

Production of dust by massive stars at high redshift

The large amounts of dust detected in sub-millimeter galaxies and quasars at high redshift pose a challenge to galaxy formation models and theories of cosmic dust formation. At z > 6 only stars of relatively high mass (> 3 Msun) are sufficiently short-lived to be potential stellar sources of dust. This review is devoted to identifying and quantifying the most important stellar channels of rapid dust formation. We ascertain the dust production efficiency of stars in the mass range 3-40 Msun using both observed and theoretical dust yields of evolved massive stars and supernovae (SNe) and provide analytical expressions for the dust production efficiencies in various scenarios. We also address the strong sensitivity of the total dust productivity to the initial mass function. From simple considerations, we find that, in the early Universe, high-mass (> 3 Msun) asymptotic giant branch stars can only be dominant dust producers if SNe generate <~ 3 x 10^-3 Msun of dust whereas SNe prevail if they are more efficient. We address the challenges in inferring dust masses and star-formation rates from observations of high-redshift galaxies. We conclude that significant SN dust production at high redshift is likely required to reproduce current dust mass estimates, possibly coupled with rapid dust grain growth in the interstellar medium.Comment: 72 pages, 9 figures, 5 tables; to be published in The Astronomy and Astrophysics Revie