Divergent effects of compounds on the hydrolysis and transpeptidation reactions of γ-glutamyl transpeptidase

A novel class of inhibitors of the enzyme γ-glutamyl transpeptidase (GGT) were evaluated. The analog OU749 was shown previously to be an uncompetitive inhibitor of the GGT transpeptidation reaction. The data in this study show that it is an equally potent uncompetitive inhibitor of the hydrolysis reaction, the primary reaction catalyzed by GGT in vivo. A series of structural analogs of OU749 were evaluated. For many of the analogs, the potency of the inhibition differed between the hydrolysis and transpeptidation reactions, providing insight into the malleability of the active site of the enzyme. Analogs with electron withdrawing groups on the benzosulfonamide ring, accelerated the hydrolysis reaction, but inhibited the transpeptidation reaction by competing with a dipeptide acceptor. Several of the OU749 analogs inhibited the transpeptidation reaction by slow onset kinetics, similar to acivicin. Further development of inhibitors of the GGT hydrolysis reaction is necessary to provide new therapeutic compounds.     

Direct shoot organogenesis from hypocotyl explants of Jatropha curcas L. an important bioenergy feedstock

Multiple shoots were induced on hypocotyl segments reared from in vitro germinated seedlings of Jatropha curcas L., using Murashige and Skoog (MS) medium supplemented with N⁶‐benzyl adenine (BA) and kinetin (kin) either alone or in combination with indole‐3‐acetic acid (IAA). The combined treatment of 7.05 μm kin and 1.425 μm IAA resulted in maximum shoot production with an average shoot bud initiation of 12.1 per explant. The regenerated microshoots were transferred to root induction medium containing half‐strength MS salts supplemented with either indole‐3‐butyric acid (IBA) or IAA. Rooting of microshoots was best achieved on half‐strength MS medium supplemented with IBA (9.8 μm ). Rooted plantlets were subsequently acclimatized under green house condition and the plantlets showed 70% survival.     

Inroads into the structure and function of intermediate filament networks

Although intermediate filaments are one of three major cytoskeletal systems of vertebrate cells, they remain the least understood with respect to their structure and function. This is due in part to the fact that they are encoded by a large gene family which is developmentally regulated in a cell and tissue type specific fashion. This article is in honor of Ueli Aebi. It highlights the studies on IF that have been carried out by our laboratory for more than 40 years. Many of our advances in understanding IF are based on conversations with Ueli which have taken place during adventurous and sometimes dangerous hiking and biking trips throughout the world.     

Whole-genome strategies for marker-assisted plant breeding

Molecular breeding for complex traits in crop plants requires understanding and manipulation of many factors influencing plant growth, development and responses to an array of biotic and abiotic stresses. Molecular marker-assisted breeding procedures can be facilitated and revolutionized through whole-genome strategies, which utilize full genome sequencing and genome-wide molecular markers to effectively address various genomic and environmental factors through a representative or complete set of genetic resources and breeding materials. These strategies are now increasingly based on understanding of specific genomic regions, genes/alleles, haplotypes, linkage disequilibrium (LD) block(s), gene networks and their contribution to specific phenotypes. Large-scale and high-density genotyping and genome-wide selection are two important components of these strategies. As components of whole-genome strategies, molecular breeding platforms and methodologies should be backed up by high throughput and precision phenotyping and e-typing (environmental assay) with strong support systems such as breeding informatics and decision support tools. Some basic strategies are discussed in this article, including (1) seed DNA-based genotyping for simplifying marker-assisted selection (MAS), reducing breeding cost and increasing scale and efficiency, (2) selective genotyping and phenotyping, combined with pooled DNA analysis, for capturing the most important contributing factors, (3) flexible genotyping systems, such as genotyping by sequencing and arraying, refined for different selection methods including MAS, marker-assisted recurrent selection and genomic selection (GS), (4) marker-trait association analysis using joint linkage and LD mapping, and (5) sequence-based strategies for marker development, allele mining, gene discovery and molecular breeding.     

Longitudinal Imaging Studies of Tumor Microenvironment in Mice Treated with the mTOR Inhibitor Rapamycin

Rapamycin is an allosteric inhibitor of mammalian target of rapamycin, and inhibits tumor growth and angiogenesis. Recent studies suggested a possibility that rapamycin renormalizes aberrant tumor vasculature and improves tumor oxygenation. The longitudinal effects of rapamycin on angiogenesis and tumor oxygenation were evaluated in murine squamous cell carcinoma (SCCVII) by electron paramagnetic resonance imaging (EPRI) and magnetic resonance imaging (MRI) to identify an optimal time after rapamycin treatment for enhanced tumor radioresponse. Rapamycin treatment was initiated on SCCVII solid tumors 8 days after implantation (500–750 mm³) and measurements of tumor pO₂ and blood volume were conducted from day 8 to 14 by EPRI/MRI. Microvessel density was evaluated over the same time period by immunohistochemical analysis. Tumor blood volume as measured by MRI significantly decreased 2 days after rapamycin treatment. Tumor pO₂ levels modestly but significantly increased 2 days after rapamycin treatment; whereas, it decreased in non-treated control tumors. Furthermore, the fraction of hypoxic area (pixels with pO₂<10 mm Hg) in the tumor region decreased 2 days after rapamycin treatments. Immunohistochemical analysis of tumor microvessel density and pericyte coverage revealed that microvessel density decreased 2 days after rapamycin treatment, but pericyte coverage did not change, similar to what was seen with anti-angiogenic agents such as sunitinib which cause vascular renormalization. Collectively, EPRI/MRI co-imaging can provide non-invasive evidence of rapamycin-induced vascular renormalization and resultant transient increase in tumor oxygenation. Improved oxygenation by rapamycin treatment provides a temporal window for anti-cancer therapies to realize enhanced response to radiotherapy.     

Estimation of the Burden of Pandemic(H1N1)2009 in Developing Countries: Experience from a Tertiary Care Center in South India

Background

The burden of the pandemic (H1N1) 2009 influenza might be underestimated if detection of the virus is mandated to diagnose infection. Using an alternate approach, we propose that a much higher pandemic burden was experienced in our institution.

Methodology/Principal Findings

Consecutive patients (n = 2588) presenting to our hospital with influenza like illness (ILI) or severe acute respiratory infection (SARI) during a 1-year period (May 2009–April 2010) were prospectively recruited and tested for influenza A by real-time RT-PCR. Analysis of weekly trends showed an 11-fold increase in patients presenting with ILI/SARI during the peak pandemic period when compared with the pre-pandemic period and a significant (P<0.001) increase in SARI admissions during the pandemic period (30±15.9 admissions/week) when compared with pre-pandemic (7±2.5) and post-pandemic periods (5±3.8). However, Influenza A was detected in less than one-third of patients with ILI/SARI [699 (27.0%)]; a majority of these (557/699, 79.7%) were Pandemic (H1N1)2009 virus [A/H1N1/09]. An A/H1N1/09 positive test was correlated with shorter symptom duration prior to presentation (p = 0.03). More ILI cases tested positive for A/H1N1/09 when compared with SARI (27.4% vs. 14.6%, P = 0.037). When the entire study population was considered, A/H1N1/09 positivity was associated with lower risk of hospitalization (p<0.0001) and ICU admission (p = 0.013) suggesting mild self-limiting illness in a majority.

Conclusion/Significance

Analysis of weekly trends of ILI/SARI suggest a higher burden of the pandemic attributable to A/H1N1/09 than estimates assessed by a positive PCR test alone. The study highlights methodological consideration in the estimation of burden of pandemic influenza in developing countries using hospital-based data that may help assess the impact of future outbreaks of respiratory illnesses.     

A novel property of DNA - as a bioflotation reagent in mineral processing

Environmental concerns regarding the use of certain chemicals in the froth flotation of minerals have led investigators to explore biological entities as potential substitutes for the reagents in vogue. Despite the fact that several microorganisms have been used for the separation of a variety of mineral systems, a detailed characterization of the biochemical molecules involved therein has not been reported so far. In this investigation, the selective flotation of sphalerite from a sphalerite-galena mineral mixture has been achieved using the cellular components of Bacillus species. The key constituent primarily responsible for the flotation of sphalerite has been identified as DNA, which functions as a bio-collector. Furthermore, using reconstitution studies, the obligatory need for the presence of non-DNA components as bio-depressants for galena has been demonstrated. A probable model involving these entities in the selective flotation of sphalerite from the mineral mixture has been discussed.     

RSARF: prediction of residue solvent accessibility from protein sequence using random forest method

Prediction of protein structure from its amino acid sequence is still a challenging problem. The complete physicochemical understanding of protein folding is essential for the accurate structure prediction. Knowledge of residue solvent accessibility gives useful insights into protein structure prediction and function prediction. In this work, we propose a random forest method, RSARF, to predict residue accessible surface area from protein sequence information. The training and testing was performed using 120 proteins containing 22006 residues. For each residue, buried and exposed state was computed using five thresholds (0%, 5%, 10%, 25%, and 50%). The prediction accuracy for 0%, 5%, 10%, 25%, and 50% thresholds are 72.9%, 78.25%, 78.12%, 77.57% and 72.07% respectively. Further, comparison of RSARF with other methods using a benchmark dataset containing 20 proteins shows that our approach is useful for prediction of residue solvent accessibility from protein sequence without using structural information. The RSARF program, datasets and supplementary data are available at http://caps.ncbs.res.in/download/pugal/RSARF/.     

Quality control genotyping for assessment of genetic identity and purity in diverse tropical maize inbred lines

Quality control (QC) genotyping is an important component in breeding, but to our knowledge there are not well established protocols for its implementation in practical breeding programs. The objectives of our study were to (a) ascertain genetic identity among 2–4 seed sources of the same inbred line, (b) evaluate the extent of genetic homogeneity within inbred lines, and (c) identify a subset of highly informative single-nucleotide polymorphism (SNP) markers for routine and low-cost QC genotyping and suggest guidelines for data interpretation. We used a total of 28 maize inbred lines to study genetic identity among different seed sources by genotyping them with 532 and 1,065 SNPs using the KASPar and GoldenGate platforms, respectively. An additional set of 544 inbred lines was used for studying genetic homogeneity. The proportion of alleles that differed between seed sources of the same inbred line varied from 0.1 to 42.3?%. Seed sources exhibiting high levels of genetic distance are mis-labeled, while those with lower levels of difference are contaminated or still segregating. Genetic homogeneity varied from 68.7 to 100?% with 71.3?% of the inbred lines considered to be homogenous. Based on the data sets obtained for a wide range of sample sizes and diverse genetic backgrounds, we recommended a subset of 50–100 SNPs for routine and low-cost QC genotyping, verified them in a different set of double haploid and inbred lines, and outlined a protocol that could be used to minimize errors in genetic analyses and breeding.