Characterisation of Inactivation Domains and Evolutionary Strata in Human X Chromosome through Markov Segmentation

Markov segmentation is a method of identifying compositionally different subsequences in a given symbolic sequence. We have applied this technique to the DNA sequence of the human X chromosome to analyze its compositional structure. The human X chromosome is known to have acquired DNA through distinct evolutionary events and is believed to be composed of five evolutionary strata. In addition, in female mammals all copies of X chromosome in excess of one are transcriptionally inactivated. The location of a gene is correlated with its ability to undergo inactivation, but correlations between evolutionary strata and inactivation domains are less clear. Our analysis provides an accurate estimate of the location of stratum boundaries and gives a high–resolution map of compositionally different regions on the X chromosome. This leads to the identification of a novel stratum, as well as segments wherein a group of genes either undergo inactivation or escape inactivation in toto. We identify oligomers that appear to be unique to inactivation domains alone

Global Metabolomic Profiling of Acute Myocarditis Caused by Trypanosoma cruzi Infection

© 2014 Gironès et al. Chagas disease is caused by Trypanosoma cruzi infection, being cardiomyopathy the more frequent manifestation. New chemotherapeutic drugs are needed but there are no good biomarkers for monitoring treatment efficacy. There is growing evidence linking immune response and metabolism in inflammatory processes and specifically in Chagas disease. Thus, some metabolites are able to enhance and/or inhibit the immune response. Metabolite levels found in the host during an ongoing infection could provide valuable information on the pathogenesis and/or identify deregulated metabolic pathway that can be potential candidates for treatment and being potential specific biomarkers of the disease. To gain more insight into those aspects in Chagas disease, we performed an unprecedented metabolomic analysis in heart and plasma of mice infected with T. cruzi. Many metabolic pathways were profoundly affected by T. cruzi infection, such as glucose uptake, sorbitol pathway, fatty acid and phospholipid synthesis that were increased in heart tissue but decreased in plasma. Tricarboxylic acid cycle was decreased in heart tissue and plasma whereas reactive oxygen species production and uric acid formation were also deeply increased in infected hearts suggesting a stressful condition in the heart. While specific metabolites allantoin, kynurenine and p-cresol sulfate, resulting from nucleotide, tryptophan and phenylalanine/tyrosine metabolism, respectively, were increased in heart tissue and also in plasma. These results provide new valuable information on the pathogenesis of acute Chagas disease, unravel several new metabolic pathways susceptible of clinical management and identify metabolites useful as potential specific biomarkers for monitoring treatment and clinical severity in patients.This work was supported by ‘‘Ministerio de Ciencia e Innovación’’ (SAF2010-17833); ‘‘Fondo de Investigaciones Sanitarias’’ (PS09/00538 and PI12/00289); ‘‘Red de Investigación de Centros de Enfermedades Tropicales’’ (RICET RD12/0018/0004); European Union (HEALTH-FE-2008-22303, ChagasEpiNet);‘‘Universidad Autónoma de Madrid’’ and ‘‘Comunidad de Madrid’’ (CC08-UAM/SAL-4440/08); AECID Cooperation with Argentine (A/025417/09 and A/031735/10), Comunidad de Madrid (S-2010/BMD-2332) and ‘‘Fundación Ramón Areces’Peer Reviewe

Evolution of complexity in the zebrafish synapse proteome

The proteome of human brain synapses is highly complex and mutated in over 130 diseases. This complexity arose from two whole genome duplications early in the vertebrate lineage. Zebrafish are used in modelling human diseases, however its synapse proteome is uncharacterised and whether the teleost-specific genome duplication (TSGD) influenced complexity is unknown. We report the characterisation of the proteomes and ultrastructure of central synapses in zebrafish and analyse the importance of the TSGD. While the TSGD increases overall synapse proteome complexity, the Post Synaptic Density (PSD) proteome of zebrafish has lower complexity than mammals. A highly conserved set of ~1000 proteins is shared across vertebrates. PSD ultrastructural features are also conserved. Lineage-specific proteome differences indicate vertebrate species evolved distinct synapse types and functions. The datasets are a resource for a wide range of studies and have important implications for the use of zebrafish in modelling human synaptic diseases

AAV Exploits Subcellular Stress Associated with Inflammation, Endoplasmic Reticulum Expansion, and Misfolded Proteins in Models of Cystic Fibrosis

Barriers to infection act at multiple levels to prevent viruses, bacteria, and parasites from commandeering host cells for their own purposes. An intriguing hypothesis is that if a cell experiences stress, such as that elicited by inflammation, endoplasmic reticulum (ER) expansion, or misfolded proteins, then subcellular barriers will be less effective at preventing viral infection. Here we have used models of cystic fibrosis (CF) to test whether subcellular stress increases susceptibility to adeno-associated virus (AAV) infection. In human airway epithelium cultured at an air/liquid interface, physiological conditions of subcellular stress and ER expansion were mimicked using supernatant from mucopurulent material derived from CF lungs. Using this inflammatory stimulus to recapitulate stress found in diseased airways, we demonstrated that AAV infection was significantly enhanced. Since over 90% of CF cases are associated with a misfolded variant of Cystic Fibrosis Transmembrane Conductance Regulator (ΔF508-CFTR), we then explored whether the presence of misfolded proteins could independently increase susceptibility to AAV infection. In these models, AAV was an order of magnitude more efficient at transducing cells expressing ΔF508-CFTR than in cells expressing wild-type CFTR. Rescue of misfolded ΔF508-CFTR under low temperature conditions restored viral transduction efficiency to that demonstrated in controls, suggesting effects related to protein misfolding were responsible for increasing susceptibility to infection. By testing other CFTR mutants, G551D, D572N, and 1410X, we have shown this phenomenon is common to other misfolded proteins and not related to loss of CFTR activity. The presence of misfolded proteins did not affect cell surface attachment of virus or influence expression levels from promoter transgene cassettes in plasmid transfection studies, indicating exploitation occurs at the level of virion trafficking or processing. Thus, we surmised that factors enlisted to process misfolded proteins such as ΔF508-CFTR in the secretory pathway also act to restrict viral infection. In line with this hypothesis, we found that AAV trafficked to the microtubule organizing center and localized near Golgi/ER transport proteins. Moreover, AAV infection efficiency could be modulated with siRNA-mediated knockdown of proteins involved in processing ΔF508-CFTR or sorting retrograde cargo from the Golgi and ER (calnexin, KDEL-R, β-COP, and PSMB3). In summary, our data support a model where AAV exploits a compromised secretory system and, importantly, underscore the gravity with which a stressed subcellular environment, under internal or external insults, can impact infection efficiency

A Model-Based Bayesian Estimation of the Rate of Evolution of VNTR Loci in Mycobacterium tuberculosis

Variable numbers of tandem repeats (VNTR) typing is widely used for studying the bacterial cause of tuberculosis. Knowledge of the rate of mutation of VNTR loci facilitates the study of the evolution and epidemiology of Mycobacterium tuberculosis. Previous studies have applied population genetic models to estimate the mutation rate, leading to estimates varying widely from around to per locus per year. Resolving this issue using more detailed models and statistical methods would lead to improved inference in the molecular epidemiology of tuberculosis. Here, we use a model-based approach that incorporates two alternative forms of a stepwise mutation process for VNTR evolution within an epidemiological model of disease transmission. Using this model in a Bayesian framework we estimate the mutation rate of VNTR in M. tuberculosis from four published data sets of VNTR profiles from Albania, Iran, Morocco and Venezuela. In the first variant, the mutation rate increases linearly with respect to repeat numbers (linear model); in the second, the mutation rate is constant across repeat numbers (constant model). We find that under the constant model, the mean mutation rate per locus is (95% CI: ,)and under the linear model, the mean mutation rate per locus per repeat unit is (95% CI: ,). These new estimates represent a high rate of mutation at VNTR loci compared to previous estimates. To compare the two models we use posterior predictive checks to ascertain which of the two models is better able to reproduce the observed data. From this procedure we find that the linear model performs better than the constant model. The general framework we use allows the possibility of extending the analysis to more complex models in the future

Reaction Chemistry and Kinetics of Corn Stalk Pyrolysis without and with Ga/HZSM-5

The bifunctional Ga/HZSM-5 catalyst has been proven having the capability to increase the selectivity of aromatics production during catalytic pyrolysis of furan and woody biomass. However, the reaction chemistry and kinetics of pyrolysis of herbaceous biomass promoted by Ga/HZSM-5 is rarely reported. Pyrolysis–gas chromatography/mass spectrometry (Py–GC/MS) analysis and non-isothermal thermogravimetric analysis at four heating rates were carried out to investigate the decomposition behavior and pyrolysis kinetics of corn stalk without and with Ga/HZSM-5. The effective activation energies for corn stalk pyrolysis were calculated by using the Friedman isoconversional method. The Py–GC/MS analysis results indicated that the Ga/HZSM-5 catalyst had a high selectivity toward producing the aromatic chemicals of xylene, toluene and benzene, whereas the major products from non-catalytic pyrolysis of corn stalk were oxygenated compounds. The presence of Ga/HZSM-5 could significantly reduce the effective activation energies of corn stalk pyrolysis from 159.9–352.4 kJ mol−1 to 41.6–99.8 kJ mol−1 in the conversion range of 0.10–0.85

Tools and techniques for solvent selection: green solvent selection guides

Driven by legislation and evolving attitudes towards environmental issues, establishing green solvents for extractions, separations, formulations and reaction chemistry has become an increasingly important area of research. Several general purpose solvent selection guides have now been published with the aim to reduce use of the most hazardous solvents. This review serves the purpose of explaining the role of these guides, highlighting their similarities and differences. How they can be used most effectively to enhance the greenness of chemical processes, particularly in laboratory organic synthesis and the pharmaceutical industry, is addressed in detail

Computational Prediction and Experimental Verification of New MAP Kinase Docking Sites and Substrates Including Gli Transcription Factors

In order to fully understand protein kinase networks, new methods are needed to identify regulators and substrates of kinases, especially for weakly expressed proteins. Here we have developed a hybrid computational search algorithm that combines machine learning and expert knowledge to identify kinase docking sites, and used this algorithm to search the human genome for novel MAP kinase substrates and regulators focused on the JNK family of MAP kinases. Predictions were tested by peptide array followed by rigorous biochemical verification with in vitro binding and kinase assays on wild-type and mutant proteins. Using this procedure, we found new ‘D-site’ class docking sites in previously known JNK substrates (hnRNP-K, PPM1J/PP2Czeta), as well as new JNK-interacting proteins (MLL4, NEIL1). Finally, we identified new D-site-dependent MAPK substrates, including the hedgehog-regulated transcription factors Gli1 and Gli3, suggesting that a direct connection between MAP kinase and hedgehog signaling may occur at the level of these key regulators. These results demonstrate that a genome-wide search for MAP kinase docking sites can be used to find new docking sites and substrates