Juvenile hormone catabolism and oviposition in the codling moth, Cydia pomonella, as functions of age, mating status, and hormone treatment.

In vitro catabolism of juvenile hormone (JH) in haemolymph of adult female Cydia pomonella was ascribed mainly to juvenile hormone esterase (JHE) activity. No significant differences were noted between virgin and mated females 0-96 h post-emergence. Changes in JHE activity did not appear dependent upon fluctuations in JH titre; conversely, changes in JHE activity could not explain the changes in JH titres. Maximal JHE activity was recorded at 24 h (331.47 +/- 47.25 pmol/h/microl; 355.93 +/- 36.68 pmol/h/microl, virgin; mated insects, respectively) and preceded the peak in JH titres at 48 h. Topical application of JH II (10 ng-10 microg) or fenoxycarb (50 ng) enhanced JHE activity up to 640 and 56%, respectively. Treatment upon emergence with 10 microg JH II induced enzymic activity for less than 24 h, and when 10 microg JH II or 50 ng fenoxycarb were applied, circulating JH titres returned to control levels within 24 h. Oviposition was highly sensitive to exogenous JH and declined significantly with dosages >100 pg. To allow a degree of oocyte maturation before JH treatment, the hormone was administered at 6, 12, 24, or 48 h post-emergence and/or females were mated. Neither measure "protected" the system; oviposition declined immediately after JH application.     

Characterizing the metabolic phenotype: a phenotype phase plane analysis.

Genome-scale metabolic maps can be reconstructed from annotated genome sequence data, biochemical literature, bioinformatic analysis, and strain-specific information. Flux-balance analysis has been useful for qualitative and quantitative analysis of metabolic reconstructions. In the past, FBA has typically been performed in one growth condition at a time, thus giving a limited view of the metabolic capabilities of a metabolic network. We have broadened the use of FBA to map the optimal metabolic flux distribution onto a single plane, which is defined by the availability of two key substrates. A finite number of qualitatively distinct patterns of metabolic pathway utilization were identified in this plane, dividing it into discrete phases. The characteristics of these distinct phases are interpreted using ratios of shadow prices in the form of isoclines. The isoclines can be used to classify the state of the metabolic network. This methodology gives rise to a "phase plane" analysis of the metabolic genotype-phenotype relation relevant for a range of growth conditions. Phenotype phase planes (PhPPs) were generated for Escherichia coli growth on two carbon sources (acetate and glucose) at all levels of oxygenation, and the resulting optimal metabolic phenotypes were studied. Supplementary information can be downloaded from our website (http://epicurus.che.udel.edu).     

Role of the C-terminal extensions of alpha-crystallins. Swapping the C-terminal extension of alpha-crystallin to alphaB-crystallin results in enhanced chaperone activity

Several small heat shock proteins contain a well conserved alpha-crystallin domain, flanked by an N-terminal domain and a C-terminal extension, both of which vary in length and sequence. The structural and functional role of the C-terminal extension of small heat shock proteins, particularly of alphaA- and alphaB-crystallins, is not well understood. We have swapped the C-terminal extensions between alphaA- and alphaB-crystallins and generated two novel chimeric proteins, alphaABc and alphaBAc. We have investigated the domain-swapped chimeras for structural and functional alterations. We have used thermal and non-thermal models of protein aggregation and found that the chimeric alphaB with the C-terminal extension of alphaA-crystallin, alphaBAc, exhibits dramatically enhanced chaperone-like activity. Interestingly, however, the chimeric alphaA with the C-terminal extension of alphaB-crystallin, alphaABc, has almost lost its activity. Pyrene solubilization and bis-1-anilino-8-naphthalenesulfonate binding studies show that alphaBAc exhibits more solvent-exposed hydrophobic pockets than alphaA, alphaB, or alphaABc. Significant tertiary structural changes are revealed by tryptophan fluorescence and near-UV CD studies upon swapping the C-terminal extensions. The far-UV CD spectrum of alphaBAc differs from that of alphaB-crystallin whereas that of alphaABc overlaps with that of alphaA-crystallin. Gel filtration chromatography shows alteration in the size of the proteins upon swapping the C-terminal extensions. Our study demonstrates that the unstructured C-terminal extensions play a crucial role in the structure and chaperone activity, in addition to generally believed electrostatic "solubilizer" function.     

The tetraspan protein epithelial membrane protein-2 interacts with beta1 integrins and regulates adhesion

The growth arrest-specific-3 (GAS3)/PMP22 proteins are members of the four-transmembrane (tetraspan) superfamily. Although the function of these proteins is poorly understood, GAS3/PMP22 proteins have been implicated in the control of growth and progression of certain cancers. Epithelial membrane protein-2 (EMP2), a GAS3/PMP22 family member, was recently identified as a putative tumor suppressor gene. Here, we addressed the normal function of EMP2 by testing the prediction that it influences integrin-related cell functions. We observed that EMP2 associates with the beta(1) integrin subunit. Co-immunoprecipitation and immunodepletion experiments indicated that approximately 60% of beta(1) integrins and EMP2 can be isolated in common protein complexes. Whereas this association between EMP2 and beta(1) integrin may be direct or indirect, it has features of integrin heterodimer selectivity. Thus, by laser confocal microscopy, EMP2 colocalized with alpha(6)beta(1) but not alpha(5)beta(1) integrin. Increased expression of EMP2 also influenced the integrin heterodimer repertoire present on the plasma membrane. EMP2 specifically increased the surface expression of the alpha(6)beta(1) integrin while decreasing that of the alpha(5)beta(1) protein. Reciprocally, reduction in EMP2 expression using a specific ribozyme decreased surface expression of alpha(6)beta(1) integrin. Accordingly, these EMP2-mediated changes resulted in a dramatic alteration in cellular adhesion to extracellular matrix proteins. This study demonstrates for the first time the interaction of a GAS3/PMP22 family member with an integrin protein and suggests that such interactions and their functional consequences are a physiologic role of GAS3/PMP22 proteins.     

Biomimetic synthesis of a water soluble conducting molecular complex of polyaniline and lignosulfonate

A new biomimetic route for the synthesis of a conducting molecular complex of polyaniline (Pani) and a natural polyelectrolyte, lignosulfonate (LGS) is presented. A poly(ethylene glycol) modified hematin (PEG-hematin) was used to catalyze the polymerization of aniline in the presence of LGS to form a Pani/LGS complex. UV-vis, FTIR, conductivity and TGA studies for the LGS-polyaniline complex indicate the presence of a thermally stable and electrically conductive form of polyaniline. Also the presence of LGS in this complex, an inexpensive byproduct from pulp processing, provides a unique combination of properties such as electronic conductivity, processability and biodegradability. The use of this conductive complex for corrosion protection is also proposed.     

Structural analysis of the maize rp1 complex reveals numerous sites and unexpected mechanisms of local rearrangement

Rp1 is a complex disease resistance locus in maize that is exceptional in both allelic variability and meiotic instability. Genomic sequence analysis of three maize BACs from the Rp1 region of the B73 inbred line revealed 4 Rp1 homologs and 18 other gene-homologous sequences, of which at least 16 are truncated. Thirteen of the truncated genes are found in three clusters, suggesting that they arose from multiple illegitimate break repairs at the same sites or from complex repairs of each of these sites with multiple unlinked DNA templates. A 43-kb region that contains an Rp1 homolog, six truncated genes, and three Opie retrotransposons was found to be duplicated in this region. This duplication is relatively recent, occurring after the insertion of the three Opie elements. The breakpoints of the duplication are outside of any genes or identified repeat sequence, suggesting a duplication mechanism that did not involve unequal recombination. A physical map and partial sequencing of the Rp1 complex indicate the presence of 15 Rp1 homologs in regions of approximately 250 and 300 kb in the B73 inbred line. Comparison of fully sequenced Rp1-homologous sequences in the region demonstrates a history of unequal recombination and diversifying selection within the Leu-rich repeat 2 region, resulting in chimeric gene structures.     

Enhanced production of antimicrobial sesquiterpenes and lipoxygenase metabolites in elicitor-treated hairy root cultures of Solanum tuberosum

Potato (Solanum tuberosum) hairy root cultures, established by infecting potato tuber discs with Agrobacterium rhizogenes, were used as a model system for the production of antimicrobial sesquiterpenes and lipoxygenase (LOX) metabolites. Of the four sesquiterpene phytoalexins (rishitin, lubimin, phytuberin and phytuberol) detected in elicitor-treated hairy root cultures, rishitin (213 g g^–1 dry wt) was the most predominant followed by lubimin (171 g g^–1 dry wt). The elicitors also induced LOX activity (25-fold increase) and LOX metabolites, mainly 9-hydroxyoctadecadienoic acid and 9-hydroxyoctadecatrienoic acid, in potato hairy root cultures. The combination of fungal elicitor plus cyclodextrin was the most effective elicitor treatment, followed by methyl jasmonate plus cyclodextrin in inducing sesquiterpenes and LOX metabolites.     

Amelioration of dextran sulfate colitis by butyrate: role of heat shock protein 70 and NF-kappaB

Butyrate enemas have been demonstrated to ameliorate inflammation in ulcerative colitis. The mechanism of this protective effect of butyrate is not known, and this study examines the effect of butyrate on epithelial function, inducible heat shock protein 70 (HSP70) expression, and NF-kappaB activation in experimental colitis. Colitis was induced in rats by oral dextran sulfate sodium (DSS) and by butyrate or saline enemas. Mucosal barrier function was assessed by electrical resistance and [14C]mannitol permeability. HSP70 production was determined by [35S]methionine labeling, Western blot analysis, and immunohistochemistry. Activation of heat shock factors (HSFs) and NF-kappaB was evaluated by EMSA. Butyrate showed a significant protection against the decrease in cell viability, increase in mucosal permeability, and polymorphonuclear neutrophil infiltration seen in DSS colitis. Butyrate inhibited HSP70 expression in DSS colitis and also inhibited the activation of HSF and NF-kappaB. Thus butyrate enema was found to be cytoprotective in DSS colitis, an effect partly mediated by suppressing activation of HSP70 and NF-kappaB.     

EPEC-activated ERK1/2 participate in inflammatory response but not tight junction barrier disruption

Enteropathogenic Escherichia coli (EPEC) alters many functions of the host intestinal epithelia. Inflammation is initiated by activation of nuclear factor (NF)-kappaB, and paracellular permeability is enhanced via a Ca2+- and myosin light-chain kinase (MLCK)-dependent pathway. The aims of this study were to identify signaling pathways by which EPEC triggers inflammation and to determine whether these pathways parallel or diverge from those that alter permeability. EPEC-induced phosphorylation and degradation of the primary inhibitor of NF-kappaB (IkappaBalpha) were tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta independent. In contrast to Salmonella typhimurium, EPEC-stimulated IkappaBalpha degradation and IL-8 expression did not require Ca2+. Instead, extracellular signal-regulated kinase (ERK)-1/2 was significantly and rapidly activated. ERK1/2 inhibitors attenuated IkappaBalpha degradation and IL-8 expression. Although ERK1/2 can activate MLCK, its inhibition had no impact on EPEC disruption of the tight junction barrier. In conclusion, EPEC-induced inflammation 1) is TNF-alpha and IL-1beta receptor independent, 2) utilizes pathways differently from S. typhimurium, 3) requires ERK1/2, and 4) employs signals that are distinct from those that alter permeability. This is the first time that EPEC-activated signaling cascades have been linked to independent functional consequences.