Mesoionic compounds with antifungal activity against Fusarium verticillioides

Background

Fungi contaminate the food of humans and animals, are a risk to health, and can cause financial losses. In this work, the antifungal activities of 16 mesoionic compounds (MI 1–16) were evaluated against mycotoxigenic fungi, including Aspergillus spp., Fusarium verticillioides and Penicillium citrinum. Furthermore, the decreased ergosterol in the total lipid content of Fusarium verticillioides was investigated.

Results

F. verticillioides was the most sensitive fungus to the mesoionic compounds. Among the evaluated compounds, MI-11 and MI-16 presented higher antifungal effects against F. verticillioides, with MIC values of 7.8 μg/ml, and MI-2 and MI-3 followed, with MICs of 15.6 μg/ml. The most active compounds were those with heterocyclic ring phenyl groups substituted by electron donor moieties (MI-11 and MI-16). Among some compounds with higher activity (MI-2, MI-11 and MI-16), decreased ergosterol content in the total lipid fraction of F. verticillioides was demonstrated. MI-2 reduced the ergosterol content approximately 40% and 80% at concentrations of 7.8 μg/ml and 15.6 μg/ml, respectively, and MI-11 and MI-16 decreased the content by 30% and 50%, respectively, when at a concentration of 7.8 μg/ml.

Conclusion

These findings indicate that mesoionic compounds have significant antifungal activity against F. verticillioides.     

ERK‐1 MAP kinase prevents TNF‐induced apoptosis through bad phosphorylation and inhibition of Bax translocation in HeLa Cells

Extracellular signal‐regulated kinase (ERK) 1/2 signaling is involved in tumor cell survival through the regulation of Bcl‐2 family members. To explore this further and to demonstrate the central role of the mitochondria in the ERK1/2 pathway we used the HeLa cellular model where apoptosis was induced by tumor necrosis factor (TNF) and cycloheximide (CHX). We show that HeLa cells overexpressing ERK‐1 displayed resistance to TNF and CHX. HeLa cells overexpressing a kinase‐deficient form of ERK‐1 (K71R) were more sensitive to TNF and CHX. In the ERK‐1 cells, Bad was phosphorylated during TNF + CHX treatment. In the HeLa wt cells and in the K71R clones TNF and CHX decreased Bad phosphorylation. ERK‐1 cells treated with TNF and CHX did not release cytochrome c from the mitochondria. By contrast, HeLa wt and K71R clones released cytochrome c. Bax did not translocate to the mitochondria in ERK‐1 cells treated with TNF + CHX. Conversely, HeLa wt and K71R clones accumulated Bax in the mitochondria. In the HeLa wt cells and in both ERK‐1 transfectants Bid was cleaved and accumulated in the mitochondria. The caspase‐8 inhibitor IETD‐FMK and the mitochondrial membrane permeabilization inhibitor bongkrekic acid (BK), partially prevented cell death by TNF + CHX. Anisomycin, a c‐Jun N‐terminal kinases activator, increased TNF‐killing. The ERK‐1 cells were resistant to TNF and anisomycin, whereas K71R clones resulted more sensitive. Our study demonstrates that in HeLa cells the ERK‐1 kinase prevents TNF + CHX apoptosis by regulating the intrinsic mitochondrial pathway through different mechanisms. Inhibition of the intrinsic pathway is sufficient to almost completely prevent cell death. J. Cell. Biochem. 108: 1166–1174, 2009. © 2009 Wiley‐Liss, Inc.     

The mechanism of covalent reaction of bromoacetyl-phenylalanyl-transfer RNA with the peptidyl-transfer RNA binding site of the Escherichia coli ribosome

Results are presented to prove that bromoacetyl-phenylalanyl-transfer RNA reacts covalently with 50 S ribosomal proteins L2 and L27 while it is bound correctly to the peptidyl site on the 70 S ribosome. Attachment of the BrAcPhe moiety to tRNA causes a 100-fold enhancement of its reactivity with ribosomes. This reactivity closely parallels binding of tRNA whether measured by poly(U) stimulation or competition with deacylated tRNA. BrAcPhe-tRNA can bind correctly to the P site as judged by puromycin releasibility and lack of tetracycline inhibition. Little significant reaction of BrAcPhe-tRNA with L2 and L27 occurs during procedures used to purify and analyze ribosomal proteins. If ribosomes are first incubated with BrAcPhe-tRNA and subsequently treated with puromycin before analysis, little inhibition of the covalent reaction with L2 and L27 is observed. In contrast, a few minor reaction products are markedly suppressed. Covalently attached BrAcPhe-tRNA is still capable of accepting an amino acid from Phe-tRNA or puromycin. The products from this reaction are found attached to proteins L2 and L27 and to a lesser extent to L15 and L16. This shows that true affinity labeling of proteins in the peptidyl binding site has been accomplished.Some covalent reaction of BrAcPhe-tRNA with the 30 S protein S18 is also observed. This reaction is not poly(U)-dependent, however, and S18-reacted BrAcPhe-tRNA is not capable of peptide bond formation with Phe-tRNA. It seems likely that reaction with S18 results from a non-functional interaction of the affinity label with the ribosome.     

Warming and drought do not influence the palatability of Quercus pubescens Willd. leaves of four European provenances

In the context of global warming, the impact of extreme drought events on trees and biotic interactions with herbivore insects is widely unknown. A faster range expansion of insects in a changing climate could lead to mass propagations of pests in forests. Therefore, the aim was to investigate the influence of climatic alterations on leaf palatability. We exposed juvenile Quercus pubescens Willd. individuals of four European provenances (Bulgaria, Germany, Hungary, and Italy) to warming and drought. In addition, we conducted a palatability experiment with the pre-exposed Q. pubescens leaves and the caterpillars of the generalist forest pest Lymantria dispar L. (gypsy moth). Consumed leaf dry material, density of trichomes, and specific leaf area were examined. Surprisingly, neither warming nor drought affected the leaf palatability, but palatability was related to the density of trichomes. The Bulgarian provenance of Q. pubescens, which had the lowest density of trichomes, was most palatable. These findings suggest that global warming and drought might not lead to more frequent infestations of the four tested European Q. pubescens provenances by L. dispar caterpillars in the future.     

ADAM12 produced by tumor cells rather than stromal cells accelerates breast tumor progression

Expression of ADAM12 is low in most normal tissues but is markedly increased in numerous human cancers, including breast carcinomas. We have previously shown that overexpression of ADAM12 accelerates tumor progression in a mouse model of breast cancer (PyMT). In this study, we found that ADAM12 deficiency reduces breast tumor progression in the PyMT model. However, the catalytic activity of ADAM12 seems to be dispensable for its tumor-promoting effect. Interestingly, we show that ADAM12 endogenously expressed in tumor-associated stroma in the PyMT model does not influence tumor progression, but that ADAM12 expression by tumor cells is necessary for tumor progression in these mice. This finding is consistent with our observation that in human breast carcinoma, ADAM12 is almost exclusively located in tumor cells and, only rarely, seen in the tumor-associated stroma. We hypothesized, however, that the tumor-associated stroma may stimulate ADAM12 expression in tumor cells, on the basis of the fact that TGF-β1 stimulates ADAM12 expression and is a well-known growth factor released from tumor-associated stroma. TGF-β1 stimulation of ADAM12-negative Lewis lung tumor cells induced ADAM12 synthesis, and growth of these cells in vivo induced more than 200-fold increase in ADAM12 expression. Our observation that ADAM12 expression is significantly higher in the terminal duct lobular units (TDLU) adjacent to human breast carcinoma compared with TDLUs found in normal breast tissue supports our hypothesis that tumor-associated stroma triggers ADAM12 expression.     

Inhibition of the Smc5/6 Complex during Meiosis Perturbs Joint Molecule Formation and Resolution without Significantly Changing Crossover or Non-crossover Levels

Meiosis is a specialized cell division used by diploid organisms to form haploid gametes for sexual reproduction. Central to this reductive division is repair of endogenous DNA double-strand breaks (DSBs) induced by the meiosis-specific enzyme Spo11. These DSBs are repaired in a process called homologous recombination using the sister chromatid or the homologous chromosome as a repair template, with the homolog being the preferred substrate during meiosis. Specific products of inter-homolog recombination, called crossovers, are essential for proper homolog segregation at the first meiotic nuclear division in budding yeast and mice. This study identifies an essential role for the conserved Structural Maintenance of Chromosomes (SMC) 5/6 protein complex during meiotic recombination in budding yeast. Meiosis-specific smc5/6 mutants experience a block in DNA segregation without hindering meiotic progression. Establishment and removal of meiotic sister chromatid cohesin are independent of functional Smc6 protein. smc6 mutants also have normal levels of DSB formation and repair. Eliminating DSBs rescues the segregation block in smc5/6 mutants, suggesting that the complex has a function during meiotic recombination. Accordingly, smc6 mutants accumulate high levels of recombination intermediates in the form of joint molecules. Many of these joint molecules are formed between sister chromatids, which is not normally observed in wild-type cells. The normal formation of crossovers in smc6 mutants supports the notion that mainly inter-sister joint molecule resolution is impaired. In addition, return-to-function studies indicate that the Smc5/6 complex performs its most important functions during joint molecule resolution without influencing crossover formation. These results suggest that the Smc5/6 complex aids primarily in the resolution of joint molecules formed outside of canonical inter-homolog pathways.