Effect of untranslated leader sequence of AMV RNA 4 and signal peptide of pathogenesis-related protein 1b on attacin gene expression, and resistance to fire blight in transgenic apple

A cDNA clone of the gene encoding attacin was used to construct three plasmid binary vectors in which attE was controlled by the cauliflower mosaic virus 35S promoter with duplicated upstream B domain (35S) (p35SAtt), 35S with the untranslated leader sequence of alfalfa mosaic virus RNA 4 (AMV) (p35SAMVAtt), and 35S with AMV and the signal peptide of pathogenesis-related protein 1b from tobacco (SP) (p35SAMVSPAtt), respectively. These plasmids and pLDB15 containing attE under the control of the potato proteinase inhibitor II (Pin2) promoter were used in Agrobacterium-mediated transformation of the apple scion cultivar `Galaxy' and the apple rootstock M.26 to enhance resistance to Erwinia amylovora, the bacterium that causes fire blight. The mean attacin content of transgenic lines containing attacin with AMV was three times higher than lines without AMV. Northern blots suggested that AMV functioned in apple as it does in other plant species by enhancing translation of attE mRNA. Transgenic `Galaxy' lines with attacin fused to SP had lower attacin content than lines without SP. In vitro assays indicated that attacin was partially degraded in the intercellular fluid of apple leaves. However, transgenic `Galaxy' lines transformed with attacin fused to SP had significantly less disease than those without SP suggesting that intercellularly secreted attacin is more effective in reducing E. amylovora infection than intracellularly localized attacin. A negative correlation was observed between attacin content and disease resistance in Pin2Att transgenic `Galaxy' lines following inoculation with E. amylovora, suggesting that attacin enhances resistance to fire blight.     

AB5 toxins: structures and inhibitor design

High-resolution crystal structures of AB₅ toxins in their native form or in complex with a variety of ligands have led to the structure-based design and discovery of inhibitors targeting different areas of the toxins. The most significant progress is the development of highly potent multivalent ligands that block binding of the toxins to their receptors.     

MIG-seq is an effective method for high-throughput genotyping in wheat (Triticum spp.)

MIG-seq (Multiplexed inter-simple sequence repeats genotyping by sequencing) has been developed as a low cost genotyping technology, although the number of polymorphisms obtained is assumed to be minimal, resulting in the low application of this technique to analyses of agricultural plants. We applied MIG-seq to 12 plant species that include various crops and investigated the relationship between genome size and the number of bases that can be stably sequenced. The genome size and the number of loci, which can be sequenced by MIG-seq, are positively correlated. This is due to the linkage between genome size and the number of simple sequence repeats (SSRs) through the genome. The applicability of MIG-seq to population structure analysis, linkage mapping, and quantitative trait loci (QTL) analysis in wheat, which has a relatively large genome, was further evaluated. The results of population structure analysis for tetraploid wheat showed the differences among collection sites and subspecies, which agreed with previous findings. Additionally, in wheat biparental mapping populations, over 3,000 SNPs/indels with low deficiency were detected using MIG-seq, and the QTL analysis was able to detect recognized flowering-related genes. These results revealed the effectiveness of MIG-seq for genomic analysis of agricultural plants with large genomes, including wheat.     

Functional expression of FIP-gts, a fungal immunomodulatory protein from Ganoderma tsugae in Sf21 insect cells

The mushrooms of diverse Lingzhi species have been traditionally consumed as luxurious functional food supplements in Chinese society. FIP-gts, a fungal immunomodulatory protein found in Song-Shan Lingzhi (Ganodera tsugae) has been proposed to possess therapeutic effects on cancer and autoimmune diseases. To produce active FIP-gts for evaluation of oral administration, a recombinant FIP-gts (rFIP-gts) fused with a 6His-tag at its C-terminus was expressed in Sf21 insect cells by the baculovirus expression system. High yield (about 70%) and purity (about 90%) of rFIP-gts was obtained by one-step nickel-affinity chromatography. The correctness of the harvested rFIP-gts was verified by Western blot and MALDI-MS analyses. Optimal expression of rFIP-gts was observed when the Sf21 cells were infected with multiplicity of infection of 10 for 72 h, and the yield was up to 47.2 microg/3 x 10(6) infected cells. The immunomodulatory activity of the purified rFIP-gts was detected as the induction of interleukin 2 released from murine splenocytes. Compared with the rFIP-gts produced in Escherichia coli cells, the rFIP-gts produced in Sf21 cells possessed evidently higher specific immunomodulatory activity.     

中国海桑属小志

本文描述中国海桑属一新种:拟海桑(Sonneratia paracaseolaris Ko, E. Y. Chen et W. Y.Chen).重新发表海桑属7种检索表,并将原隶属海桑组的海南海桑S.hainanensis Ko, E. Y.Chenet W.Y.Chen改隶无瓣海桑组,因其过去被认为是花瓣的部分实为退化雄蕊.     

Mutagenicity and polycyclic aromatic hydrocarbon content of fumes from heated cooking oils produced in Taiwan

According to epidemiologic studies, exposure of women to fumes from cooking oils appears to be an important risk factor for lung cancer. Fume samples from three different commercial cooking oils frequently used in Taiwan were collected and analyzed for mutagenicity in the Salmonella/microsome assay. Polycyclic aromatic hydrocarbons were extracted from the samples and identified by HPLC chromatography. Extracts from three cooking oil fumes were found to be mutagenic in the presence of S9 mix. All samples contained dibenz[a,h]anthracene (DB[a,h]A) and benz[a]anthracene (B[a]A). Concentration of DB[a,h]A and B[a]A were 1.9 and 2.2 μg/m³ in fumes from lard oil, 2.1 and 2.3 μg/m³ in soybean oil, 1.8 and 1.3 μg/m³ in peanut oil, respectively. Benzo[a]pyrene (B[a]P) was identified in fume samples of soybean and peanut oil, in concentrations of 19.6 and 18.3 μg/m³, in this order. These results provide experimental evidence and support the findings of epidemiologic observations, in which women exposed to the emitted fumes of cooking oils are at increased risk of contracting lung cancer.     

Granulysin, a T cell product, kills bacteria by altering membrane permeability

Granulysin, a protein located in the acidic granules of human NK cells and cytotoxic T cells, has antimicrobial activity against a broad spectrum of microbial pathogens. A predicted model generated from the nuclear magnetic resonance structure of a related protein, NK lysin, suggested that granulysin contains a four alpha helical bundle motif, with the alpha helices enriched for positively charged amino acids, including arginine and lysine residues. Denaturation of the polypeptide reduced the alpha helical content from 49 to 18% resulted in complete inhibition of antimicrobial activity. Chemical modification of the arginine, but not the lysine, residues also blocked the antimicrobial activity and interfered with the ability of granulysin to adhere to Escherichia coli and Mycobacterium tuberculosis. Granulysin increased the permeability of bacterial membranes, as judged by its ability to allow access of cytosolic ss-galactosidase to its impermeant substrate. By electron microscopy, granulysin triggered fluid accumulation in the periplasm of M. tuberculosis, consistent with osmotic perturbation. These data suggest that the ability of granulysin to kill microbial pathogens is dependent on direct interaction with the microbial cell wall and/or membrane, leading to increased permeability and lysis.