Enantiomeric Composition of the trans-Dihydrodiols Produced from Phenanthrene by Fungi

The trans-dihydrodiols produced during the metabolism of phenanthrene by Cunninghamella elegans, Syncephalastrum racemosum, and Phanerochaete chrysosporium were purified by high-performance liquid chromatography (HPLC). The enantiomeric compositions and optical purities of the trans-dihydrodiols were determined to compare interspecific differences in the regio- and stereoselectivity of the fungal enzymes. Circular dichroism spectra of the trans-dihydrodiols were obtained, and the enantiomeric composition of each preparation was analyzed by HPLC with a chiral stationary-phase column. The phenanthrene trans-1,2-dihydrodiol produced by C. elegans was a mixture of the 1R,2R and 1S,2S enantiomers in variable proportions. The phenanthrene trans-3,4-dihydrodiol produced by P. chrysosporium was the optically pure 3R,4R enantiomer, but that produced by S. racemosum was a 68:32 mixture of the 3R,4R and 3S,4S enantiomers. The phenanthrene trans-9,10-dihydrodiol produced by P. chrysosporium was predominantly the 9S,10S enantiomer, but those produced by C. elegans and S. racemosum were predominantly the 9R,10R enantiomer. The results indicate that although different fungi may exhibit similar regioselectivity, there still may be differences in stereoselectivity that depend on the species and the cultural conditions.     

Comparison of the primary structure ofwaxy proteins (granule-bound starch synthase) between polyploid wheats and related diploid species

Thewaxy proteins encoded by the genomes A, B, and D in polyploid wheats and related diploid species were isolated by SDS-PAGE. The N-terminal amino acid sequences of mature proteins and V8 protease-induced fragments were determined. A total of five amino acid substitutions was detected in these sequences, which represent about 10% of the whole sequences of thewaxy proteins. A comparison of these sequences in polyploid wheats with those in related diploid species revealed the following: (i)waxy proteins encoded by the A genome of polyploid wheats were identical to that ofTriticum monococcum, (ii) thewaxy protein encoded by the B genome ofT. turgidum was identical to that ofT. searsii, but differed from those ofT. speltoides andT. longissimum by one amino acid substitution, (iii) thewaxy protein encoded by the B genome ofT. aestivum differed from that encoded by the B genome ofT. turgidum by one amino acid substitution, and (iv) thewaxy protein encoded by the D genome ofT. aestivum was identical to that ofT. tauschii.     

Xiphidorus amazonensis n. sp. (Nematoda: Longidoridae) from the Brazilian Amazon Basin

Xiphidorus amazonensis n. sp. was found in the rhizospheres of Jatropha curcas, Musa sp., Anona muricata, Cassia tora, Panicum laxum, Paspalum fasciculatum, Aeschynomene sensitiva, Saccharum officinarum, Manihot esculenta, Abelmoschus esculentus, Tamarindus indica, Mangifera indica, Vigna unguiculata, Zea mays, Commelina sp., Cyperus rotundus, Fimbristylis miliacea, Citrus sinensis, and Eichhornia crassipes on the Amazon River island of Xiborena, approximately 40 km southeast of Manaus, capital of the State of Amazonas. The type habitat is flooded annually for about 6 months by the Amazon River. Xiphidorus amazonensis n. sp. differs from the closely related species Xiphidorus yepesara Monteiro, 1976 by the larger size, by a, b, and c values, and by the rounded tail terminus. It also resembles Xiphidorus tucumanensis Chaves and Coomans, 1984, but can be distinguished by its larger size, larger a, b, and c values, more conical female tail, bilobed amphidial pouch, and the presence of a spermatheca full of sperm.     

The tipuloid dipterans (Diptera,Tipulidae, Limoniidae) from the Putorana Plateau,with a description of Dactylolabis tschernovi sp. n.

The first data are presented on the tipuloid dipterans from the Putorana Plateau, the Central Siberian Plateau: the crane-flies, Tipula (Arctotipula) oklandi Al., Tipula (Pterelachisus) tundrensis stackelbergiana Lack., Tipula (Vestiplex) arctica Curt., and Tipula (Yamatotipula) lionota Holm.; and the limoniids, Dactylolabis (Dactylolabis) novaezemblae (Al.) and Dactylolabis (Dactylolabis) tschernovi sp. n. A new species, Dactylolabis (D.) tschernovi sp. n., is described from the adult males and females. The male of the new species is very similar to those of Dactylolabis (Dactylolabis) carbonaria Sav. and Dactylolabis (Dactylolabis) satanas Sav. but can be distinguished from the former by the presence of a distinct throat, by the coloration of the head, femora, tibiae, and abdominal segments, as well as by the spear-shaped interbase of the hypopygium; and from the latter species, by the fully developed wings and by the absence of large spines at the base of the interbases.     

Comparison of Pratylenchus penetrans Infection and Maladera castanea Feeding on Strawberry Root Rot

The interaction of lesion nematodes, black root rot disease caused by Rhizoctonia fragariae, and root damage caused by feeding of the scarab larva, Maladera castanea, was determined in greenhouse studies. Averaged over all experiments after 12 weeks, root weight was reduced 13% by R. fragariae and 20% by M. castanea. The percentage of the root system affected by root rot was increased by inoculation with either R. fragariae (35% more disease) or P. penetrans (50% more disease) but was unaffected by M. castanea. Rhizoctonia fragariae was isolated from 9.2% of the root segments from plants not inoculated with R. fragariae. The percentage of R. fragariae-infected root segments was increased 3.6-fold by inoculation with R. fragariae on rye seeds. The presence of P. penetrans also increased R. fragariae root infection. The type of injury to root systems was important in determining whether roots were invaded by R. fragariae and increased the severity of black root rot. Pratylenchus penetrans increased R. fragariae infection and the severity of black root rot. Traumatic cutting action by Asiatic garden beetle did not increase root infection or root disease by R. fragariae. Both insects and diseases need to be managed to extend the productive life of perennial strawberry plantings.     

Hemolysis induced by Bacillus cereus sphingomyelinase

Bacillus cereus sphingomyelinase (Bc-SMase) induces hemolysis of sheep erythrocytes which contain large amounts of sphingomyelin. We investigated the mechanism of this hemolysis in comparison to that induced by Clostridium perfringens alpha-toxin. Pertussis toxin, a Gi-specific inhibitor, N-oleoylethernolamine, a ceramidase inhibitor, and dihydrosphingosine, a sphingosine kinase inhibitor, did not inhibit the hemolysis by Bc-SMase, but did inhibit that by alpha-toxin. Bc-SMase broadly bound to whole membranes, and alpha-toxin specifically bound to the detergent-resistant membrane fractions, lipid rafts. The level of ceramide production induced by Bc-SMase in sheep erythrocytes was 6- to 15-fold that induced by alpha-toxin, when the extent of the hemolysis by Bc-SMase was the same as that by the toxin. However, the level of ceramide production induced by Bc-SMase in SM-liposomes was equal to that triggered by the toxin, when the carboxyl fluorescein-release from liposomes induced by Bc-SMase was the same as that induced by alpha-toxin. Confocal laser microscopy showed that treatment of the cells with Bc-SMase resulted in the formation of ceramide-rich domains. A photobleaching analysis suggested that treatment of the cells with Bc-SMase leads to a reduction in membrane fluidity. These results show that Bc-SMase-induced hemolysis of sheep erythrocytes is related to the formation of interface between ceramide-rich domains and ceramide-poor domains through production of ceramide from SM.     

Attenuating l-lysine production by deletion of ddh and lysE and their effect on l-threonine and l-isoleucine production in Corynebacterium glutamicum

The fermentative production of l-threonine and l-isoleucine with Corynebacterium glutamicum is usually accompanied by the by-production of l-lysine, which shares partial biosynthesis pathway with l-threonine and l-isoleucine. Since the direct precursor for l-lysine synthesis, diaminopimelate, is a component of peptidoglycan and thus essential for cell wall synthesis, reducing l-lysine by-production could be troublesome. Here, a basal strain with eliminated l-lysine production was constructed from the wild type C. glutamicum ATCC13869 by deleting the chromosomal ddh and lysE. Furthermore, the basal strain as well as the ddh single mutant strain was engineered for l-threonine production by over-expressing lysC1, hom1 and thrB, and for l-isoleucine production by over-expressing lysC1, hom1, thrB and ilvA1. Fermentation experiments with the engineered strains showed that (i) deletion of ddh improved l-threonine production by 17%, and additional deletion of lysE further improved l-threonine production by 28%; (ii) deletion of ddh improved l-isoleucine production by 8% and improved cell growth by 21%, whereas additional deletion of lysE had no further influence on both l-isoleucine production and cell growth; (iii) l-lysine by-production was reduced by 95% and 86% in l-threonine and l-isoleucine production, respectively, by deletion of ddh and lysE. This is the first report on improving l-threonine and l-isoleucine production by deleting ddh and lysE in C. glutamicum. The results demonstrate deletion of ddh and lysE as an effective strategy to reduce l-lysine by-production without surrendering the cell growth of C. glutamicum.     

Initial predation and parasitism by muricid whelks demonstrated by the correspondence between drilled holes and their apparent enveloper

The morphologies of drilled holes enveloped by three coexisting muricid whelks, Morula musiva, Cronia margariticola and Ergalatax contractus, were compared. The results were used to demonstrate the replacement of feeders using drilled holes, and to estimate the frequency of this occurrence in the field. In the laboratory, M. musiva made circular internal holes, whereas C. margariticola and E. contractus made oval holes, when they preyed upon the mussel Hormomya mutabilis. The maximum inner diameter of the hole correlated with the driller's shell height in all three muricid species. In the field, mussels enveloped by M. musiva had circular drilled holes, as was found in the laboratory, and the correlation between maximum diameter and enveloper's shell height was not significantly different from that found in the laboratory. In contrast, holes enveloped by C. margariticola in the field were more circular than those observed in the laboratory, and there was no correlation between maximum diameter and enveloper's shell height. Statistical analyses of these results indicated that more than half of the mussels being ingested by C. margariticola had been drilled by M. musiva, i.e. C. margariticola would often kleptoparasitize or scavenge M. musiva's prey by taking over the drilled hole. Few M. musiva would take over the holes drilled by C. margariticola or E. contractus. Furthermore, probably few M. musiva would take over the hole made by a conspecific initial drilling predator. It appears that E. contractus rarely initiates predation by drilling.     

Blockade by N-methylhydroxylamine or activation of guanylate cyclase and elevations of guanosine 3′,5′-monophosphate levels in nervous tissues

Hydroxylamine and N-methylhydroxylamine prevented the activation of soluble guanylate cyclase by the endogenous activator as well as by nitroso compounds such as N-methyl-N′-nitro-N-nitroguanidine or nitroprusside, while the other derivaties of hydroxylamine were ineffective. Hydroxylamine and N-methylhydroxylamine did not alter the basal guanylate cyclase activity of purified enzyme preparations. Kinetics analysis indicated that N-methylhydroxylamine competes with N-methyl-N′nitro-N-nitrosuguanidine for guanylate cyclase. The activation of guanylate cyclase by N-methyl-N′-nitro-N-nitrosoguanidine and its inhibition by N-methylhydroxylamine were reversible reactions. These efects of N-methyl-N′-nitro-N-nitrosoguanine and N-methylhydroxylamine were observed with guanylate cyclase from other tissues.N-Methylhydroxylamine preveneed the increase of guanosine 3′,5′-monophosphate (cyclic GMP) levels in cerebellar slices of guinea pig by N-methyl-N′-nitro-N-nitroguanidine, veratridine and adenosine, while the elevalations of adenosine 3′,5′-monophosphate by these agents were not affected. N-Methylhyroxylamine also blocked the increased of cyclic GMP levels by carbachol, prostaglandin E₁ and N-methyl-N′-nitro-N-nitrosoguanidine in neuroblastoma N1E 115 cells. Thus N-methylhydroxylamine prevents the activation of guanylate cyclase and the increased synthesis of cyclic GMP in responses to transmitters without blocking the synthesis of cyclic GMP via basal enzyme activity.     

Response of the zoophytophagous predators Macrolophus pygmaeus and Nesidiocoris tenuis to volatiles of uninfested plants and to plants infested by prey or conspecifics

Knowledge about the orientation mechanisms used by two important predaceous mirids (Macrolophus pygmaeus Rambour and Nesidiocoris tenuis (Reuter)) in finding their prey (whitefly Bemisia tabaci (Gennadius) and the tomato borer Tuta absoluta (Meyrick)) is limited. In a Y-tube olfactometer, we tested the behavioral responses of naïve and experienced predators to uninfested plants, herbivore-induced plant volatiles (HIPVs) from plants infested with T. absoluta and/or B. tabaci, the sex pheromone of T. absoluta, and volatiles produced by plants injured by the predators. Nesidiocoris tenuis responds to volatiles produced by uninfested plants only after experience with the plant, whereas naïve and experienced M. pygmaeus show positive chemotaxis. Both predators are attracted to volatiles from prey-infested plants, and we provide the first evidence that experience affects this response in M. pygmaeus. Infestation of the same plant by both prey species elicited similar responses by the two predators as plants infested by either herbivore singly. Neither predator responded to sex pheromones of T. absoluta. Macrolophus pygmaeus avoided plants injured by conspecifics, while N. tenuis females were attracted by such plants. The implications of these results for augmentative biological control are discussed.     

rpoD Gene Pyrosequencing for the Assessment of Pseudomonas Diversity in a Water Sample from the Woluwe River

A water sample from a noncontaminated site at the source of the Woluwe River (Belgium) was analyzed by culture-dependent and -independent methods. Pseudomonas isolates were identified by sequencing and analysis of the rpoD gene. Culture-independent methods consisted of cloning and pyrosequencing of a Pseudomonas rpoD amplicon from total DNA extracted from the same sample and amplified with selective rpoD gene primers. Among a total of 14,540 reads, 6,228 corresponded to Pseudomonas rpoD gene sequences by a BLAST analysis in the NCBI database. The selection criteria for the reads were sequences longer than 400 bp, an average Q₄₀ value greater than 25, and >85% identity with a Pseudomonas species. Of the 6,228 Pseudomonas rpoD sequences, 5,345 sequences met the established criteria for selection. Sequences were clustered by phylogenetic analysis and by use of the QIIME software package. Representative sequences of each cluster were assigned by BLAST analysis to a known Pseudomonas species when the identity with the type strain was greater than or equal to 96%. Twenty-six species distributed among 12 phylogenetic groups or subgroups within the genus were detected by pyrosequencing. Pseudomonas stutzeri, P. moraviensis, and P. simiae were the only cultured species not detected by pyrosequencing. The predominant phylogenetic group within the Pseudomonas genus was the P. fluorescens group, as determined by culture-dependent and -independent analyses. In all analyses, a high number of putative novel phylospecies was found: 10 were identified in the cultured strains and 246 were detected by pyrosequencing, indicating that the diversity of Pseudomonas species has not been fully described.     

Assimilation of E. coli cells by Daphnia magna on the whole organism level

Consumption of E. coli cells by Daphnia magna was studied. It was found that this organism not only ingested E. coli cells but digested them as demonstrated by the release of ¹⁴CO₂ originating from E. coli grown on ¹⁴C-glucose, and by the transfer of the radioactive label from parental Daphnia to their progenies. In addition the effect of antibiotics on the consumption of E. coli cells by Daphnia magna was studied. In long incubation times, antibiotics inhibited bacterial uptake by Daphnia. The microflora isolated from Daphnia was found to be capable of causing leakage of enzymes out of E. coli cells thus playing at least a partial role in the digestion of E. coli cells by Daphnia.     

p27Kip1 Mediates Addiction of Ovarian Cancer Cells to MYCC (c-MYC) and Their Dependence on MYC Paralogs

The MYCC (c-MYC) gene is amplified in 30–60% of human ovarian cancers. We assessed the functional significance of MYCC amplification by siRNA inhibition of MYCC or MYC paralogs in a panel of ovarian cancer cell lines expressing varying levels of MYCC. Inactivation of MYCC inhibited cell proliferation and induced replicative senescence only in lines with amplified MYCC, indicating that these cells are addicted to continued MYCC overexpression. In contrast, siRNA knockdown of all three MYC isoforms inhibited proliferation of MYCC non-amplified ovarian cancer cells without inducing replicative senescence, and did not inhibit the proliferation of telomerase-immortalized ovarian surface epithelial cells. The arrest induced by MYCC knockdown was accompanied by an increase in the level of the Cdk inhibitor p27^Kip1 and a decrease in cyclin A expression and Cdk2 activity, and could be reversed by RNAi knockdown of p27^Kip1 or Rb, or by overexpression of cyclin A/Cdk2. The arrest induced by knockdown of all three MYC isoforms could similarly be reversed by p27^Kip1 knockdown. Our findings indicate that the addiction of MYCC-amplified ovarian cancer cells to MYCC differs from the dependence of MYCC non-amplified cancer cells on MYC paralogs, but both are mediated, at least in part, by p27^Kip1. They also suggest that growth of ovarian cancers may be blocked by inhibition of MYCC or MYC paralogs.     

两栖动物性别决定相关基因的研究进展

两栖动物的性别决定机制主要包括遗传性别决定(genetic sex determination,GSD)和环境性别决定(environmental sex determination,ESD).近年来,在两栖动物性别决定和性腺分化机制的研究中,运用分子生物学技术探讨性别决定相关基因及其相互关系方面的研究已获得新的成果....     

A spatially dynamic cohort of regulatory genes in the endomesodermal gene network of the sea urchin embryo

A gene regulatory network subcircuit comprising the otx, wnt8, and blimp1 genes accounts for a moving torus of gene expression that sweeps concentrically across the vegetal domain of the sea urchin embryo. Here we confirm by mutation the inputs into the blimp1cis-regulatory module predicted by network analysis. Its essential design feature is that it includes both activation and autorepression sites. The wnt8 gene is functionally linked into the subcircuit in that cells receiving this ligand generate a β-catenin/Tcf input required for blimp1 expression, while the wnt8 gene in turn requires a Blimp1 input. Their torus-like spatial expression patterns and gene regulatory analysis indicate that the genes even-skipped and hox11/13b are also entrained by this subcircuit. We verify the cis-regulatory inputs of even-skipped predicted by network analysis. These include activation by β-catenin/Tcf and Blimp1, repression within the torus by Hox11/13b, and repression outside the torus by Tcf in the absence of Wnt8 signal input. Thus even-skipped and hox11/13b, along with blimp1 and wnt8, are members of a cohort of torus genes with similar regulatory inputs and similar, though slightly out-of-phase, expression patterns, which reflect differences in cis-regulatory design.     

Sister Chromatid Exchange Frequencies in Escherichia coli Analyzed by Recombination at the dif Resolvase Site

Sister chromatid exchange (SCE) in Escherichia coli results in the formation of circular dimer chromosomes, which are converted back to monomers by a compensating exchange at the dif resolvase site. Recombination at dif is site specific and can be monitored by utilizing a density label assay that we recently described. To characterize factors affecting SCE frequency, we analyzed dimer resolution at the dif site in a variety of genetic backgrounds and conditions. Recombination at dif was increased by known hyperrecombinogenic mutations such as polA, dut, and uvrD. It was also increased by a fur mutation, which increased oxidative DNA damage. Recombination at dif was eliminated by a recA mutation, reflecting the role of RecA in SCE and virtually all homologous recombination in E. coli. Interestingly, recombination at dif was reduced to approximately half of the wild-type levels by single mutations in either recB or recF, and it was virtually eliminated when both mutations were present. This result demonstrates the importance of both RecBCD and RecF to chromosomal recombination events in wild-type cells.     

Isolation and structural analyses of positional isomers of 61,6m-di-O-α-d-mannopyranosyl-cyclomaltooctaose (m=2–5) and 6-O-α-(n-O-α-d-mannopyranosyl)-α-d-mannopyranosyl-cyclomaltooctaose (n=2, 3, 4, and 6)

Eight positional isomers of 6¹,6^m-di-O-α-d-mannopyranosyl-cyclomaltooctaose (γCD) (m=2–5) and 6-O-α-(n-O-α-d-mannopyranosyl)-d-mannopyranosyl-γCD (n=2, 3, 4, and 6) in a mixture of products from γCD and d-mannose by condensation reaction of α-mannosidase from jack bean were isolated by HPLC. The structures of four isomers of 6-O-α-(n-O-α-d-mannopyranosyl)-d-mannopyranosyl-γCD were elucidated by NMR spectroscopy. On the other hand, four positional isomers of 6¹,6^m-di-O-α-d-mannopyranosyl-γCD were determined by LC–MS analysis of degree of polymerization of the branched oligosaccharides produced by enzymatic degradation with bacterial saccharifying α-amylase (BSA), and combination of BSA and glucoamylase. Similarly cyclomaltodextrin glucanotransferase also digested these isomers.