Biosynthesis and characterization of [15N]actinomycin D and conformational analysis by nitrogen-15 nuclear magnetic resonance

We describe the production and characterization of actinomycin D labeled with 15N at all twelve nitrogen positions. Cultures of Streptomyces parvulus were incubated in the presence of racemic [15N]glutamic acid and, following an initial delay, labeled antibiotic was produced. Evidence is presented that the D enantiomorph of glutamic acid was ultimately used for actinomycin biosynthesis. The 15N NMR spectrum at 10.14 and 20.47 MHz of the labeled drug in CDCl3 is presented. All nitrogens except the phenoxazone chromophore nitrogen are inverted when spectra are obtained under broad-band proton irradiation conditions. All 15N resonances have been assigned, and the proton-nitrogen one-bond coupling constants were determined in CDCl3 to be 92.5 +/- 0.3 Hz for the valine and threonine amide protons by both 1H and 15N NMR. 15N NMR spectra were also obtained in dimethyl sulfoxide, methanol, and water in order to probe solvent interactions with the peptide nitrogens and carbonyl groups. Large downfield shifts (greater than 5 ppm) were seen for the Pro, sarcosine, and methylvaline resonances when the solvent was changed from dimethyl sulfoxide to water. Smaller downfield shifts were observed for the Val and Thr peaks. These results are discussed in terms of a model for the solution conformation of the actinomycin pentapeptide rings based on different hydrogen-bonding interactions in the monomer in organic solvents and the dimer which is formed in water.     

Microbiome dynamics of human epidermis following skin barrier disruption

Background

Recent advances in sequencing technologies have enabled metagenomic analyses of many human body sites. Several studies have catalogued the composition of bacterial communities of the surface of human skin, mostly under static conditions in healthy volunteers. Skin injury will disturb the cutaneous homeostasis of the host tissue and its commensal microbiota, but the dynamics of this process have not been studied before. Here we analyzed the microbiota of the surface layer and the deeper layers of the stratum corneum of normal skin, and we investigated the dynamics of recolonization of skin microbiota following skin barrier disruption by tape stripping as a model of superficial injury.

Results

We observed gender differences in microbiota composition and showed that bacteria are not uniformly distributed in the stratum corneum. Phylogenetic distance analysis was employed to follow microbiota development during recolonization of injured skin. Surprisingly, the developing neo-microbiome at day 14 was more similar to that of the deeper stratum corneum layers than to the initial surface microbiome. In addition, we also observed variation in the host response towards superficial injury as assessed by the induction of antimicrobial protein expression in epidermal keratinocytes.

Conclusions

We suggest that the microbiome of the deeper layers, rather than that of the superficial skin layer, may be regarded as the host indigenous microbiome. Characterization of the skin microbiome under dynamic conditions, and the ensuing response of the microbial community and host tissue, will shed further light on the complex interaction between resident bacteria and epidermis.     

Respect for outsiders? respect for the law? the moral evaluation of high scale issues by US immigration officers

High-scale morality is the study of moral ideas and sentiments deployed in relations that encompass multiple, geographically or socially distant populaces. The envisioning of distant people, their attributed moral personhood, the evaluation of their perceived behaviour, and the rectification of wrongs through the use of powerful organizations are key topics in high-scale morality. High-scale morality differs from existing anthropological approaches that emphasize local ethnography or contrastive moral ideas; it addresses the moralization of issues like world hunger, the drug trade, or international migration. The officers of the US Immigration and Naturalization Service understand and evaluate legal and illegal immigrants, as well as directly enacting moral rectification for the US polity. As they resolve moral dilemmas on their job, they utilize pervasive models for moral thought and action in capitalist, individualist, stratified, and bureaucratized societies. The article finishes by considering directions in which anthropology can contribute to understanding the moral dimension of global issues.     

Genetic dissection of Al tolerance QTLs in the maize genome by high density SNP scan

Background

Aluminum (Al) toxicity is an important limitation to food security in tropical and subtropical regions. High Al saturation on acid soils limits root development, reducing water and nutrient uptake. In addition to naturally occurring acid soils, agricultural practices may decrease soil pH, leading to yield losses due to Al toxicity. Elucidating the genetic and molecular mechanisms underlying maize Al tolerance is expected to accelerate the development of Al-tolerant cultivars.

Results

Five genomic regions were significantly associated with Al tolerance, using 54,455 SNP markers in a recombinant inbred line population derived from Cateto Al237. Candidate genes co-localized with Al tolerance QTLs were further investigated. Near-isogenic lines (NILs) developed for ZmMATE2 were as Al-sensitive as the recurrent line, indicating that this candidate gene was not responsible for the Al tolerance QTL on chromosome 5, qALT5. However, ZmNrat1, a maize homolog to OsNrat1, which encodes an Al³⁺ specific transporter previously implicated in rice Al tolerance, was mapped at ~40 Mbp from qALT5. We demonstrate for the first time that ZmNrat1 is preferentially expressed in maize root tips and is up-regulated by Al, similarly to OsNrat1 in rice, suggesting a role of this gene in maize Al tolerance. The strongest-effect QTL was mapped on chromosome 6 (qALT6), within a 0.5 Mbp region where three copies of the Al tolerance gene, ZmMATE1, were found in tandem configuration. qALT6 was shown to increase Al tolerance in maize; the qALT6-NILs carrying three copies of ZmMATE1 exhibited a two-fold increase in Al tolerance, and higher expression of ZmMATE1 compared to the Al sensitive recurrent parent. Interestingly, a new source of Al tolerance via ZmMATE1 was identified in a Brazilian elite line that showed high expression of ZmMATE1 but carries a single copy of ZmMATE1.

Conclusions

High ZmMATE1 expression, controlled either by three copies of the target gene or by an unknown molecular mechanism, is responsible for Al tolerance mediated by qALT6. As Al tolerant alleles at qALT6 are rare in maize, marker-assisted introgression of this QTL is an important strategy to improve maize adaptation to acid soils worldwide.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-153) contains supplementary material, which is available to authorized users.     

31P and 1H NMR studies of ethanolamine-linked phosphoglycerides metabolism in human T lymphocytes.

Aqueous and organic extracts of peripheral human T lymphocytes and of T lymphoblastoid cell lines have been examinated by 31P and 1H NMR spectroscopy in order to study the metabolism of ethanolamine (Etn) linked phosphoglycerides. The results show that the Etn concentration in the culture medium determines the composition of Etn-containing metabolites and phospholipids. The effect of phorbol esters, stimulating the synthesis and the breakdown of choline-containing phospholipids has been also studied. A phorbol 12-myristate 13-acetate (PMA) dependent membrane phosphatidylethanolamine hydrolysis, presumably mediated by protein kinase C activity, has been demonstrated.     

Mapping of citrullinated fibrinogen B-cell epitopes in rheumatoid arthritis by imaging surface plasmon resonance

Introduction  

Rheumatoid arthritis (RA) frequently involves the loss of tolerance to citrullinated antigens, which may play a role in pathogenicity. Citrullinated fibrinogen is commonly found in inflamed synovial tissue and is a frequent target of autoantibodies in RA patients. To obtain insight into the B-cell response to citrullinated fibrinogen in RA, its autoepitopes were systematically mapped using a new methodology.     

Canonical Wnt signaling: high-throughput RNAi widens the path

The canonical Wnt signaling pathway is highly conserved in evolution, widely used throughout animal development, and frequently hyperactive in cancer. Although Wnt signaling has been the subject of extensive genetic analysis in the past, some 200 genes have now been identified as candidate modulators of this pathway by a recent study using high-throughput RNAi screening.     

Tacrine derivatives-acetylcholinesterase interaction: 1H NMR relaxation study

Two acetylcholinesterase (AChE) inhibitors structurally related to Tacrine, 6-methoxytacrine (1a) and 9-heptylamino-6-methoxytacrine (1b), and their interaction with Electrophorus Electricus AChE were investigated. The complete assignment of the 1H and 13C NMR spectra of 1a and 1b was performed by mono-dimensional and homo- and hetero-correlated two-dimensional NMR experiments. This study was undertaken to elucidate the interaction modes between AChE and 1a and 1b in solution, using NMR. The interaction between the two inhibitors and AChE was studied by the analysis of the motional parameters non-selective and selective spin-lattice relaxation times, thereby allowing the motional state of 1a and 1b, both free and bound with AChE, to be defined. The relaxation data pointed out the ligands molecular moiety most involved in the binding with AChE. The relevant ligand/enzyme interaction constants were also evaluated for both compounds and resulted to be 859 and 5412M(-1) for 1a and1b, respectively.     

Quantification of global transcription patterns in prokaryotes using spotted microarrays

We describe an analysis, applicable to any spotted microarray dataset produced using genomic DNA as a reference, that quantifies prokaryotic levels of mRNA on a genome-wide scale. Applying this to Mycobacterium tuberculosis, we validate the technique, show a correlation between level of expression and biological importance, define the complement of invariant genes and analyze absolute levels of expression by functional class to develop ways of understanding an organism's biology without comparison to another growth condition.