Flower-specific expression of <Emphasis Type="Italic">Arabidopsis</Emphasis><Emphasis Type="Italic">PCS1</Emphasis> driven by <Emphasis Type="Italic">AGAMOUS</Emphasis> second intron in tobacco decreases the fertility of transgenic plants

The PROMOTION OF CELL SURVIVAL 1 (PCS1) gene, encoding an aspartic protease, has an important role in determining the fate of cells in embryonic development and reproduction processes in Arabidopsis. To explore the potential function of the PCS1 gene in generating reproductive sterility, we placed the PCS1 gene under the control of an 1,869-bp nucleotide sequence from the 3′ end of the second intron (AG-I) of Arabidopsis AGAMOUS and CaMV 35S (–60) minimal promoter [AG-I-35S (–60)::PCS1], and introduced it into tobacco. RT–PCR results demonstrated that the PCS1 gene driven by AG-I-35S (–60) chimeric promoter was expressed only in anthers and carpels in the reproductive tissues of transgenic tobacco. Compared to wild-type plants, all AG-I-35S (–60) and AG-I-35S (–60)::PCS1 transgenic lines showed a normal phenotype throughout the vegetative growth phase. However, during the reproductive stage, most AG-I-35S (–60)::PCS1 transgenic plant anthers displayed delayed dehiscence, failed dehiscence, petalody and hypoplasia, and the pollen grains had different shapes and sizes with a distorted, shrunken, or collapsed morphology. Moreover, three transgenic lines, PCS1-1, PCS1-3 and PCS1-4, showed higher sterility than wild-type and AG-I-35S (–60) transgenic plants, respectively. These results showed that the construct of AG-I-35S (–60)::PCS1 was partially effective at preventing seed set and provided a novel sterility strategy.     

In vitro deamidation of human triosephosphate isomerase

The effects of pH, temperature, buffer ion, ionic strength, protein concentration, and substrate on the rates of specific, spontaneous deamidations of Asn-15 and Asn-71 of human triosephosphate isomerase were examined. Elevated temperature and pH facilitate the deamidations, and the deamidation rate is dependent on the specific buffer ions indicating a general base catalysis mechanism. The presence of substrate also enhances the rates of deamidation. The effect of substrate may be related to conformational changes in the catalytic center which are known to cause changes in the subunit-subunit contact sites where Asn-15 and Asn-71 are located. The enhanced deamidation in the presence of substrate may, in part, account for the more rapid rate of deamidation observed in vivo.     

Xenoestrogenic short ethoxy chain nonylphenol is oxidized by a flavoprotein alcohol dehydrogenase from <Emphasis Type="Italic">Ensifer</Emphasis> sp. strain AS08

The ethoxy chains of short ethoxy chain nonylphenol (NPEO_av2.0, containing average 2.0 ethoxy units) were dehydrogenated by cell-free extracts from Ensifer sp. strain AS08 grown on a basal medium supplemented with NPEO_av2.0. The reaction was coupled with the reduction in 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide and phenazine methosulfate. The enzyme (NPEO_av2.0 dehydrogenase; NPEO-DH) was purified to homogeneity with a yield of 20% and a 56-fold increase in specific activity. The molecular mass of the native enzyme was 120 kDa, consisting of two identical monomer units (60 kDa). The gene encoding NPEO-DH was cloned, which consisted of 1,659 bp, corresponding to a protein of 553 amino acid residues. The deduced amino acid sequence agreed with the N-terminal amino acid sequence of the purified NPEO-DH. The presence of a flavin adenine dinucleotide (FAD)-binding motif and glucose–methanol–choline (GMC) oxidoreductase signature motifs strongly suggested that the enzyme belongs to the GMC oxidoreductase family. The protein exhibited homology (40–45% identity) with several polyethylene glycol dehydrogenases (PEG-DHs) of this family, but the identity was lower than those (approximately 58%) among known PEG-DHs. The substrate-binding domain was more hydrophobic compared with those of glucose oxidase and PEG-DHs. The recombinant protein had the same molecular mass as the purified NPEO-DH and dehydrogenated PEG400-2000, NPEO_av2.0 and its components, and NPEOav10, but only slight or no activity was found using diethylene glycol, triethylene glycol, and PEG200. English edition: The paper was edited by a native speaker through American Journal Experts ().     

A Franco-Italian investigation of funerary rituals in the Roman world, “les rites et la mort à Pompéi”, the plant part: a preliminary report

This paper deals with the botanical study of a family funerary enclosure located in the Porta Nocera necropolis in Pompeii (southwestern Italy). This study is part of a Franco-Italian programme investigating Roman funerary rituals. The choice of the context was due to the exceptional preservation of the archaeological features, which offered the opportunity to observe the remains of the proceedings which took place in a funerary enclosure in great detail. An adequate methodology had to be developed and a 3D recording of every single artefact or ecofact has been made. Both ground surfaces and graves provided botanical results. Those from the ground surfaces consisted mainly of fruit offering residues (especially fig and grape), while a much wider range of species was observed in the tombs, including cereals, pulses, other kinds of fruits, weeds and bread/pastry. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Lipoxin A-induced inhibition of human natural killer cell cytotoxicity: studies on stereospecificity of inhibition and mode of action

Human leukocyte-derived lipoxin A (LXA; 5S,-6R,15S-trihydroxy-7,9,13-trans-11-cis-eicosatetraenoic acid) inhibits the cytotoxic activity of human natural killer (NK) cells. LXA and three of its isomers were prepared by total organic synthesis and assayed for activity with human NK cells. Dose-response studies showed that biologically derived LXA and synthetic LXA were equally effective in inhibiting NK cell cytotoxicity. 6S-LXA, with its 6S-OH group in an (S) configuration, proved to be approximately half as potent as LXA. In contrast, 6S-11-trans-LXA and 11-trans-LXA displayed virtually no inhibitory activities. The methyl esters of both LXA and 6S-LXA proved to be more potent than their corresponding free acids. Thus, LXA inhibition of NK cells displays clear-cut stereochemistry. In the absence of putative inhibitors, NK cells bind to their targets to form conjugates. This event is followed by polarization of the NK Golgi apparatus, which moves towards the plasma membrane that is in contact with the target cell. However, in the presence of either the methyl ester or free acid of LXA, the Golgi apparati of NK cells bound to their targets were randomly oriented. In contrast, neither 6S-11-trans-LXA nor the potent NK inhibitor prostaglandin E2 affected the polarization. Furthermore, although prostaglandin E2 resulted in a decrease in NK-target cell binding efficiency, LXA and its isomers failed to affect conjugate formation. Together these results indicate that LXA-induced inhibition of NK cytotoxicity does not act on NK cell binding but may block cytotoxicity by disrupting "signals" involved in the specific orientation of the Golgi. Thus, this latter event may appear to be important in cytotoxicity.     

Human erythrocyte flickering: temperature,ATP concentration,water transport,and cell aging,plus a computer simulation

Images of human erythrocytes from a healthy donor were recorded under differential interference contrast (DIC) microscopy; they were acquired rapidly (~336 Hz) and the intensity of the centermost pixel of each cell was recorded for ~60 s (20,000 values). Various techniques were used to analyze the data, including detrended fluctuation analysis (DFA) and multiscale entropy (MSE); however, power spectrum analysis was deemed the most appropriate for metrifying and comparing results. This analysis was used to compare cells from young and old populations, and after perturbing normal conditions, with changes in temperature, adenosine triphosphate (ATP) concentration (using NaF, an inhibitor of glycolysis, and α-toxin, a pore-forming molecule used to permeabilize red cells to ATP), and water transport rates [using glycerol, and p-chloromercuriphenylsulfonic acid (pCMBS) to inhibit aquaporins, AQPs]. There were measurable differences in the membrane fluctuation characteristics in populations of young and old cells, but there was no significant change in the flickering time series on changing the temperature of an individual cell, by depleting it of ATP, or by competing with the minor water exchange pathway via AQP3 using glycerol. However, pCMBS, which inhibits AQP1, the major water exchange pathway, inhibited flickering in all cells, and yet it was restored by the membrane intercalating species dibutyl phthalate (DBP). We developed a computer model to simulate acquired displacement spectral time courses and to evaluate various methods of data analysis, and showed how the flexibility of the membrane, as defined in the model, affects the flickering time course.     

Competition between n-alkane-assimilating yeasts and bacteria during colonization of sandy soil microcosms

An n-alkane-assimilating strain of Candida tropicalis was selected in sandy soil inoculated with microorganisms from contaminated sites. Competition experiments with n-alkane utilizers from different strain collections confirmed that yeasts overgrow bacteria in sandy soil. Acidification of the soil is one of the colonization factors useful for the yeasts. It can be counteracted by addition of bentonite, a clay mineral with high ion exchange capacity, but not, however, by kaolin. Strains of different yeast species showed different levels of competitiveness. Strains of Arxula adeninivorans, Candida maltosa, and Yarrowia lipolytica overgrew strains of C. tropicalis, C. shehatae or Pichia stipitis. Two strains of C. maltosa and Y. lipolytica coexisted during several serial transfers under microcosm conditions. Received: 20 October 1999 / Received revision: 26 January 2000 / Accepted: 27 January 2000     

Countrywide Survey for MERS-Coronavirus Antibodies in Dromedaries and Humans in Pakistan

Middle East Respiratory Syndrome Coronavirus (MERS-CoV) is a zoonotic pathogen capable of causing severe respiratory disease in humans. Although dromedary camels are considered as a major reservoir host, the MERS-CoV infection dynamics in camels are not fully understood. Through surveillance in Pakistan, nasal (n = 776) and serum (n = 1050)samples were collected from camels between November 2015 and February 2018. Samples were collected from animal markets, free-roaming herds and abattoirs. An in-house ELISA was developed to detect IgG against MERS-CoV. A total of 794 camels were found seropositive for MERS-CoV. Prevalence increased with the age and the highest seroprevalence was recorded in camels aged [ 10 years (81.37%) followed by those aged 3.1–10 years (78.65%) and B 3 years (58.19%).Higher prevalence was observed in female (78.13%) as compared to male (70.70%). Of the camel nasal swabs, 22 were found to be positive by RT-qPCR though with high Ct values. Moreover, 2,409 human serum samples were also collected from four provinces of Pakistan during 2016–2017. Among the sampled population, 840 humans were camel herders.Although we found a high rate of MERS-CoV antibody positive dromedaries (75.62%) in Pakistan, no neutralizing antibodies were detected in humans with and without contact to camels.