Enantioselective Uptake and Translocation of a Novel Chiral Neonicotinoid Insecticide Cycloxaprid in Youdonger (Brassica campestris subsp. Chinensis)

For a novel potential commercial chiral pesticide, an independent study on the fate characteristics and residues of each stereoisomer is essential if the application rates for the pesticide and human exposure are to be reduced. The absorption and translocation behavior of a chiral insecticide, cycloxaprid, in plants treated by root immersion and blade smearing was studied using ¹⁴C‐labeling tracer techniques. With the root treatment, total absorption of (1R;8S)‐cycloxaprid (RS) (12.39%) was much greater than that of (1S;8R)‐cycloxaprid (SR) (3.31%) at 192 h after treatment (HAT). The mass concentrations ( RS / SR ) of cycloxaprid in the roots, cotyledons, leaf 1, leaf 2, and leaf 3 were 37.0/16.8, 8.3/2.8, 11.7/6.5, 5.1/4.8, and 8.0/4.7 mg kg^‐1 (fresh weight), respectively, at 192 HAT at an initial concentration 1.6 mg kg^‐1. With the foliar application treatment, no significant difference was observed between the total absorption of RS (3.11%) and SR (4.03%) at the end of the treatment. Both acropetal and basipetal transport of absorbed ¹⁴C occurred and more than 71.83% of absorbed RS and 82.42% of SR remained in the treated leaf. Stereoselective absorption was observed during root uptake but not during foliar absorption. Chirality 25:686–691, 2013. © 2013 Wiley Periodicals, Inc.     

CmPEX6, a Gene Involved in Peroxisome Biogenesis,Is Essential for Parasitism and Conidiation by the Sclerotial Parasite Coniothyrium minitans

Coniothyrium minitans is a sclerotial parasite of the plant-pathogenic fungus Sclerotinia sclerotiorum, and conidial production and parasitism are two important aspects for commercialization of this biological control agent. To understand the mechanism of conidiation and parasitism at the molecular level, we constructed a transfer DNA (tDNA) insertional library with the wild-type strain ZS-1. A conidiation-deficient mutant, ZS-1TN22803, was uncovered through screening of this library. This mutant could produce pycnidia on potato dextrose agar (PDA), but most were immature and did not bear conidia. Moreover, this mutant lost the ability to parasitize or rot the sclerotia of S. sclerotiorum. Analysis of the tDNA flanking sequences revealed that a peroxisome biogenesis factor 6 (PEX6) homolog of Saccharomyces cerevisiae, named CmPEX6, was disrupted by the tDNA insertion in this mutant. Targeted gene replacement and gene complementation tests confirmed that a null mutation of CmPEX6 was responsible for the phenotype of ZS-1TN22803. Further analysis showed that both ZS-1TN22803 and the targeted replacement mutants could not grow on PDA medium containing oleic acid, and they produced much less nitric oxide (NO) and hydrogen peroxide (H₂O₂) than wild-type strain ZS-1. The conidiation of ZS-1TN22803 was partially restored by adding acetyl-CoA or glyoxylic acid to the growth media. Our results suggest that fatty acid β-oxidation, reactive oxygen and nitrogen species, and possibly other unknown pathways in peroxisomes are involved in conidiation and parasitism by C. minitans.     

Knockdown of dual specificity phosphatase 4 enhances the chemosensitivity of MCF-7 and MCF-7/ADR breast cancer cells to doxorubicin

BackgroundBreast cancer is the major cause of cancer-related deaths in females world-wide. Doxorubicin-based therapy has limited efficacy in breast cancer due to drug resistance, which has been shown to be associated with the epithelial-to-mesenchymal transition (EMT). However, the molecular mechanisms linking the EMT and drug resistance in breast cancer cells remain unclear. Dual specificity phosphatase 4 (DUSP4), a member of the dual specificity phosphatase family, is associated with cellular proliferation and differentiation; however, its role in breast cancer progression is controversial.MethodsWe used cell viability assays, Western blotting and immunofluorescent staining, combined with siRNA interference, to evaluate chemoresistance and the EMT in MCF-7 and adriamycin-resistant MCF-7/ADR breast cancer cells, and investigate the underlying mechanisms.ResultsKnockdown of DUSP4 significantly increased the chemosensitivity of MCF-7 and MCF-7/ADR breast cancer cells to doxorubicin, and MCF-7/ADR cells which expressed high levels of DUSP4 had a mesenchymal phenotype. Furthermore, knockdown of DUSP4 reversed the EMT in MCF-7/ADR cells, as demonstrated by upregulation of epithelial biomarkers and downregulation of mesenchymal biomarkers, and also increased the chemosensitivity of MCF-7/ADR cells to doxorubicin.ConclusionsDUSP4 might represent a potential drug target for inhibiting drug resistance and regulating the process of the EMT during the treatment of breast cancer.     

Comparative characterization of novel ene‐reductases from cyanobacteria

The growing importance of biocatalysis in the syntheses of enantiopure molecules results from the benefits of enzymes regarding selectivity and specificity of the reaction and ecological issues of the process. Ene‐reductases (ERs) from the old yellow enzyme family have received much attention in the last years. These flavo‐enzymes catalyze the trans‐specific reduction of activated C?C bonds, which is an important reaction in asymmetric synthesis, because up to two stereogenic centers can be created in one reaction. However, limitations of ERs described in the literature such as their moderate catalytic activity and their strong preference for NADPH promote the search for novel ERs with improved properties. In this study, we characterized nine novel ERs from cyanobacterial strains belonging to different taxonomic orders and habitats. ERs were identified with activities towards a broad spectrum of alkenes. The reduction of maleimide was catalyzed with activities of up to 35.5 U mg^?1 using NADPH. Ketoisophorone and (R)‐carvone, which were converted to the highly valuable compounds (R)‐levodione and (2R,5R)‐dihydrocarvone, were reduced with reaction rates of up to 2.2 U mg^?1 with NADPH. In contrast to other homologous ERs from the literature, NADH was accepted at moderate to high rates as well: Enzyme activities of up to 16.7 U mg^?1 were obtained for maleimide and up to 1.3 U mg^?1 for ketoisophorone and (R)‐carvone. Additionally, excellent stereoselectivities were achieved in the reduction of (R)‐carvone (97–99% de). In particular, AnabaenaER3 from Anabaena variabilis ATCC 29413 and AcaryoER1 from Acaryochloris marina MBIC 11017 were identified as useful biocatalysts. Therefore, novel ERs from cyanobacteria with high catalytic efficiency were added to the toolbox for the asymmetric reduction of alkenes. Biotechnol. Bioeng. 2013; 110: 1293–1301. © 2012 Wiley Periodicals, Inc.     

Synthesis and biological evaluation of 18F-labled 2-phenylindole derivatives as PET imaging probes for β-amyloid plaques

A novel series of fluorinated 2-phenylindole derivatives were synthesized and evaluated as β-amyloid imaging probes for PET. The in vitro inhibition assay demonstrated that their binding affinities for Aβ_1–42 aggregates ranged from 28.4 to 1097.8 nM. One ligand was labeled with ¹⁸F ([¹⁸F]1a) for its high affinity (K_i = 28.4 nM), which was also confirmed by in vitro autoradiography experiments on brain sections of transgenic mouse (C57BL6, APPswe/PSEN1, 11 months old, male). In vivo biodistribution experiments in normal mice showed that this radiotracer displayed high initial uptake (5.82 ± 0.51% ID/g at 2 min) into and moderate washout (2.77 ± 0.31% ID/g at 60 min) from the brain. [¹⁸F]1a could be developed as a promising new PET imaging probe for Aβ plaques although necessary modifications are still needed.     

XBP-1u suppresses autophagy by promoting the degradation of FoxO1 in cancer cells

Autophagy is activated to maintain cellular energy homeostasis in response to nutrient starvation. However, autophagy is not persistently activated, which is poorly understood at a mechanistic level. Here, we report that turnover of FoxO1 is involved in the dynamic autophagic process caused by glutamine starvation. X-box-binding protein-1u (XBP-1u) has a critical role in FoxO1 degradation by recruiting FoxO1 to the 20S proteasome. In addition, the phosphorylation of XBP-1u by extracellular regulated protein kinases1/2 (ERK1/2) on Ser61 and Ser176 was found to be critical for the increased interaction between XBP-1u and FoxO1 upon glutamine starvation. Furthermore, knockdown of XBP-1u caused the sustained level of FoxO1 and the persistent activation of autophagy, leading to a significant decrease in cell viability. Finally, the inverse correlation between XBP-1u and FoxO1 expression agrees well with the expression profiles observed in many human cancer tissues. Thus, our findings link the dynamic process of autophagy to XBP-1u-induced FoxO1 degradation.     

Global Changes in the Mammary Epigenome Are Induced by Hormonal Cues and Coordinated by Ezh2

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Highlights? The mammary epigenome is highly sensitive to steroid hormones at specific developmental stages ? Ezh2 links hormonal cues to changes in chromatin structure and gene expression ? Progesterone is a key in vivo regulator of Ezh2 ? Hormone-induced chromatin changes likely play a role in the initiation of breast cancer     

Massively Parallel Sequencing Reveals the Complex Structure of an Irradiated Human Chromosome on a Mouse Background in the Tc1 Model of Down Syndrome

Down syndrome (DS) is caused by trisomy of chromosome 21 (Hsa21) and presents a complex phenotype that arises from abnormal dosage of genes on this chromosome. However, the individual dosage-sensitive genes underlying each phenotype remain largely unknown. To help dissect genotype – phenotype correlations in this complex syndrome, the first fully transchromosomic mouse model, the Tc1 mouse, which carries a copy of human chromosome 21 was produced in 2005. The Tc1 strain is trisomic for the majority of genes that cause phenotypes associated with DS, and this freely available mouse strain has become used widely to study DS, the effects of gene dosage abnormalities, and the effect on the basic biology of cells when a mouse carries a freely segregating human chromosome. Tc1 mice were created by a process that included irradiation microcell-mediated chromosome transfer of Hsa21 into recipient mouse embryonic stem cells. Here, the combination of next generation sequencing, array-CGH and fluorescence in situ hybridization technologies has enabled us to identify unsuspected rearrangements of Hsa21 in this mouse model; revealing one deletion, six duplications and more than 25 de novo structural rearrangements. Our study is not only essential for informing functional studies of the Tc1 mouse but also (1) presents for the first time a detailed sequence analysis of the effects of gamma radiation on an entire human chromosome, which gives some mechanistic insight into the effects of radiation damage on DNA, and (2) overcomes specific technical difficulties of assaying a human chromosome on a mouse background where highly conserved sequences may confound the analysis. Sequence data generated in this study is deposited in the ENA database, Study Accession number: ERP000439.