Double-stranded RNA-mediated gene silencing of cysteine proteases (falcipain-1 and -2) of Plasmodium falciparum

Malaria remains a public health problem of enormous magnitude, affecting over 500 million people every year. Lack of success in the past in the development of new drug/vaccines has mainly been attributed to poor understanding of the functions of different parasite proteins. Recently, RNA interference (RNAi) has emerged as a simple and incisive technique to study gene functions in a variety of organisms. In this study, we report the results of RNAi by double-stranded RNA of cysteine protease genes (falcipain-1 and -2) in the malaria parasite, Plasmodium falciparum. Using RNAi directed towards falcipain genes, we demonstrate that blocking the expression of these genes results in severe morphological abnormalities in parasites, inhibition of parasite growth in vitro and substantial accumulation of haemoglobin in the parasite. The inhibitory effects produced by falcipain double-stranded (ds)RNAs are reminiscent of the effects observed upon administering E-64, a cysteine protease inhibitor. The parasites treated with falcipain's dsRNAs also show marked reduction in the levels of corresponding endogenous falcipain mRNAs. We also demonstrate that dsRNAs of falcipains are broken into short interference RNAs approximately 25 nucleotides in size, a characteristic of RNAi, which in turn activates sequence-specific nuclease activity in the malaria parasites. These results thus provide more evidence for the existence of RNAi in P. falciparum and also suggest possibilities for using RNAi as an effective tool to determine the functions of the genes identified from the P. falciparum genome sequencing project.     

WU-Blast2 server at the European Bioinformatics Institute

Since 1995, the WU-BLAST programs (http://blast.wustl.edu) have provided a fast, flexible and reliable method for similarity searching of biological sequence databases. The software is in use at many locales and web sites. The European Bioinformatics Institute's WU-Blast2 (http://www.ebi.ac.uk/blast2/) server has been providing free access to these search services since 1997 and today supports many features that both enhance the usability and expand on the scope of the software.     

Evaluation of a new modulus mapping technique to investigate microstructural features of human teeth

Teeth contain several calcified tissues with junctions that provide interfaces between dissimilar tissues. These junctions have been difficult to characterize because of their small size. In this work a new technique using a combination of atomic force microscopy (AFM) and a force-displacement transducer was used to simultaneously study the surface topography and map mechanical properties of the junctions and adjacent hard tissues. Prepared specimens from human third molars were scanned by an AFM piezo-tube in contact mode. To measure the dynamic viscoelastic properties of the material a small sinusoidal force was superimposed on the contact force and the resulting displacement amplitude and the phase shift between the force and amplitude were measured. This force modulation technique was used to map the local variation of nanomechanical properties of intertubular dentin, peritubular dentin, enamel, dentin-enamel junction (DEJ) and peritubular-intertubular dentin junction (PIJ). This new technique allowed us to measure the widths of these junctions in addition to local variation in dentin and enamel without causing plastic deformation to the material and with 2 orders of magnitude increase in spatial resolution compared with previous studies that used discrete nanoindentation techniques. Due to the ability to analyze the sample line-by-line, the distribution functions associated with the width of the DEJ and PIJ were conveniently obtained for specific intratooth locations. The data suggested, for three third molar specimens, a DEJ width of 2-3 microm with full-width half-maximum (FWHM) of 0.7 microm and PIJ width of 0.5-1.0 microm with 0.3 microm FWHM. The intertubular dentin storage modulus variation was between 17 and 23 GPa with a mean value of 21 GPa. The range of storage modulus for enamel near the DEJ was between 51 and 74 GPa with a mean value of 63 GPa.     

Cloning and characterization of three fatty alcohol oxidase genes from Candida tropicalis strain ATCC 20336

Candida tropicalis (ATCC 20336) converts fatty acids to long-chain dicarboxylic acids via a pathway that includes among other reactions the oxidation of omega-hydroxy fatty acids to omega-aldehydes by a fatty alcohol oxidase (FAO). Three FAO genes (one gene designated FAO1 and two putative allelic genes designated FAO2a and FAO2b), have been cloned and sequenced from this strain. A comparison of the DNA sequence homology and derived amino acid sequence homology between these three genes and previously published Candida FAO genes indicates that FAO1 and FAO2 are distinct genes. Both genes were individually cloned and expressed in Escherichia coli. The substrate specificity and K(m) values for the recombinant FAO1 and FAO2 were significantly different. Particularly striking is the fact that FAO1 oxidizes omega-hydroxy fatty acids but not 2-alkanols, whereas FAO2 oxidizes 2-alkanols but not omega-hydroxy fatty acids. Analysis of extracts of strain H5343 during growth on fatty acids indicated that only FAO1 was highly induced under these conditions. FAO2 contains one CTG codon, which codes for serine (amino acid 177) in C. tropicalis but codes for leucine in E. coli. An FAO2a construct, with a TCG codon (codes for serine in E. coli) substituted for the CTG codon, was prepared and expressed in E. coli. Neither the substrate specificity nor the K(m) values for the FAO2a variant with a serine at position 177 were radically different from those of the variant with a leucine at that position.     

Intrathecal baclofen withdrawal syndrome- a life-threatening complication of baclofen pump: A case report

Background  

Intrathecal baclofen pump has been used effectively with increasing frequency in patients with severe spasticity, particularly for those patients who are unresponsive to conservative pharmacotherapy or develop intolerable side effects at therapeutic doses of oral baclofen. Drowsiness, nausea, headache, muscle weakness, light-headedness and return of pretreatment spasticity can be caused by intrathecal pump delivering an incorrect dose of baclofen. Intrathecal baclofen withdrawal syndrome is a very rare, potentially life-threatening complication of baclofen pump caused by an abrupt cessation of intrathecal baclofen.     

Expression of multiple forms of polygalacturonase gene during ripening in banana fruit.

The activity of polygalacturonase (PG, E.C 3.2.1.15) during ripening in climacteric fruits has been positively correlated with softening of the fruit tissue and differential expression of its gene is suspected to be regulated by the plant hormone ethylene. We have cloned four partial cDNAs, MAPG1 (acc. no. AF311881), MAPG2 (acc. no. AF311882), MAPG3 (acc. no. AF542382) and MAPG4 (acc. no. AY603341) for PG genes and studied their differential expression during ripening in banana. MAPG3 and MAPG4 are believed to be ripening related and regulated by ethylene whereas MAPG2 is associated more with senescence. MAPG1 shows constitutive expression and is not significantly expressed in fruit tissue. The genomic clone MAGPG (acc. No. AY603340) includes the complete MAPG3 gene, which consists of four exons and three introns. The structure of the gene has more similarity to tomato abscission PG rather than tomato fruit PG. It is concluded that softening during ripening in banana fruit results from the concerted action of at least four PG genes, which are differentially expressed during ripening.     

Diversity of Prokaryotic Communities Indigenous to Acid Mine Drainage and Related Rocks from Baiyin Open-Pit Copper Mine Stope,China

Metagenomic approach permits us to obtain the latent resources from culturable and unculturable microorganisms in ecosystem. In this study, high-throughput sequencing was practiced to comprehensively probe prokaryotic community within extreme acidic environment of Baiyin open-pit mine stope, which varied in pH and other physicochemical parameters. Bioinformatics analysis was further accomplished to process millions of Illumina reads and analyzed alpha and beta diversities, and prokaryotic community profile in different samples obtained from the acidic mine stope. Diversity indices such as ACE, Chao, Shannon, and Simpson were varied among samples. Both taxon richness and evenness were significantly higher in the solid samples than that of the water samples. Taxonomic diversity was unexpectedly higher within confined pit ecosystem. Most of the sequences were assigned to phyla Proteobacteria, Firmicutes, and Acidobacteria. In archaea, Euryarchaeota and Thaumarchaeota were major phyla reported, however, archaea occupied very little share in the metagenome. At class level, variation in community structure was higher within samples. Among iron- and sulfur-related acidophiles, 30.8% of the sequences were unidentified at genera level, while the remaining were dominated by sulfur and/or iron oxidizing Acidithiobacillus and heterotrophic Acidiphilum related groups. The community profile of solid and water groups was different and metagenomic biomarkers were higher in solid, while acidophiles and archaea were reported only in water group by using LEfSe. Among samples, community structure and abundance was varied in terms of OTUs abundance, which clearly indicates spatial variation and proposed the influence of physicochemical and geochemical properties on phylogenetic diversity. This study offers numerous treasured datasets for better understanding the community composition under the influence of geochemical and physicochemical factors and possible novelty in terms of taxonomic/phylogenetic diversity in acidic ecosystem.     

Meta-QTL analysis of seed iron and zinc concentration and content in common bean (<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis> L.)

Key message

Twelve meta-QTL for seed Fe and Zn concentration and/or content were identified from 87 QTL originating from seven population grown in sixteen field trials. These meta-QTL include 2 specific to iron, 2 specific to zinc and 8 that co-localize for iron and zinc concentrations and/or content.

Abstract

Common bean (Phaseolus vulgaris L.) is the most important legume for human consumption worldwide and it is an important source of microelements, especially iron and zinc. Bean biofortification breeding programs develop new varieties with high levels of Fe and Zn targeted for countries with human micronutrient deficiencies. Biofortification efforts thus far have relied on phenotypic selection of raw seed mineral concentrations in advanced generations. While numerous quantitative trait loci (QTL) studies have been conducted to identify genomic regions associated with increased Fe and Zn concentration in seeds, these results have yet to be employed for marker-assisted breeding. The objective of this study was to conduct a meta-analysis from seven QTL studies in Andean and Middle American intra- and inter-gene pool populations to identify the regions in the genome that control the Fe and Zn levels in seeds. Two meta-QTL specific to Fe and two meta-QTL specific to Zn were identified. Additionally, eight Meta QTL that co-localized for Fe and Zn concentration and/or content were identified across seven chromosomes. The Fe and Zn shared meta-QTL could be useful candidates for marker-assisted breeding to simultaneously increase seed Fe and Zn. The physical positions for 12 individual meta-QTL were identified and within five of the meta-QTL, candidate genes were identified from six gene families that have been associated with transport of iron and zinc in plants.

     

Mapping of novel salt tolerance QTL in an Excalibur × Kukri doubled haploid wheat population

Key message

Novel QTL for salinity tolerance traits have been detected using non-destructive and destructive phenotyping in bread wheat and were shown to be linked to improvements in yield in saline fields.

Abstract

Soil salinity is a major limitation to cereal production. Breeding new salt-tolerant cultivars has the potential to improve cereal crop yields. In this study, a doubled haploid bread wheat mapping population, derived from the bi-parental cross of Excalibur?×?Kukri, was grown in a glasshouse under control and salinity treatments and evaluated using high-throughput non-destructive imaging technology. Quantitative trait locus (QTL) analysis of this population detected multiple QTL under salt and control treatments. Of these, six QTL were detected in the salt treatment including one for maintenance of shoot growth under salinity (QG_(1–5).asl-7A), one for leaf Na⁺ exclusion (QNa.asl-7A) and four for leaf K⁺ accumulation (QK.asl-2B.1, QK.asl-2B.2, QK.asl-5A and QK:Na.asl-6A). The beneficial allele for QG_(1–5).asl-7A (the maintenance of shoot growth under salinity) was present in six out of 44 mainly Australian bread and durum wheat cultivars. The effect of each QTL allele on grain yield was tested in a range of salinity concentrations at three field sites across 2 years. In six out of nine field trials with different levels of salinity stress, lines with alleles for Na⁺ exclusion and/or K⁺ maintenance at three QTL (QNa.asl-7A, QK.asl-2B.2 and QK:Na.asl-6A) excluded more Na⁺ or accumulated more K⁺ compared to lines without these alleles. Importantly, the QK.asl-2B.2 allele for higher K⁺ accumulation was found to be associated with higher grain yield at all field sites. Several alleles at other QTL were associated with higher grain yields at selected field sites.

     

Corrigendum to “Synthesis,structure-activity relationship and molecular docking studies of 3-O-flavonol glycosides as cholinesterase inhibitors” [Bioorg. Med. Chem. 26 (12) (2018) 3696–3706]

1. Download high-res image (135KB)
2. Download full-size image