Information Transfer in Gonadotropin-releasing Hormone (GnRH) Signaling: EXTRACELLULAR SIGNAL-REGULATED KINASE (ERK)-MEDIATED FEEDBACK LOOPS CONTROL HORMONE SENSING*

Cell signaling pathways are noisy communication channels, and statistical measures derived from information theory can be used to quantify the information they transfer. Here we use single cell signaling measures to calculate mutual information as a measure of information transfer via gonadotropin-releasing hormone (GnRH) receptors (GnRHR) to extracellular signal-regulated kinase (ERK) or nuclear factor of activated T-cells (NFAT). This revealed mutual information values <1 bit, implying that individual GnRH-responsive cells cannot unambiguously differentiate even two equally probable input concentrations. Addressing possible mechanisms for mitigation of information loss, we focused on the ERK pathway and developed a stochastic activation model incorporating negative feedback and constitutive activity. Model simulations revealed interplay between fast (min) and slow (min-h) negative feedback loops with maximal information transfer at intermediate feedback levels. Consistent with this, experiments revealed that reducing negative feedback (by expressing catalytically inactive ERK2) and increasing negative feedback (by Egr1-driven expression of dual-specificity phosphatase 5 (DUSP5)) both reduced information transfer from GnRHR to ERK. It was also reduced by blocking protein synthesis (to prevent GnRH from increasing DUSP expression) but did not differ for different GnRHRs that do or do not undergo rapid homologous desensitization. Thus, the first statistical measures of information transfer via these receptors reveals that individual cells are unreliable sensors of GnRH concentration and that this reliability is maximal at intermediate levels of ERK-mediated negative feedback but is not influenced by receptor desensitization.     

Polyamine Content in Relation to Ploidy Level and to Different Organs of Nicotiana plumbaginifolia

Levels of free and bound polyamines (PAs) have been analysedin different organs of diploid, haploid and hypohaploid phenotypesof Nicotiana plumbaginifolia. In leaves and stems there is apositive correlation between the content of the PAs bound tomacromolecules and ploidy level. In contrast, in flowers a negativecorrelation is found with bound spermidine (Spd) levels. Besidesthe ploidy degree, the high levels of Spd in the flowers ofthe haploid and hypohaploid phenotypes might be also due tothe abnormal floral development and thus supports the view thatthis triamine could play a role in floral differentiation. (Received January 16, 1990; Accepted June 6, 1990)     

<Emphasis Type="Italic">Escherichia coli</Emphasis> modular coculture system for resveratrol glucosides production

In bio-based fermentation, the overall bioprocess efficiency is significantly affected by the metabolic burden associated with the expression of complete biosynthetic pathway as well as precursor and cofactor generating enzymes into a single microbial cell. To attenuate such burden by compartmentalizing the enzyme expression, recently synthetic biologists have used coculture or poly-culture techniques for biomolecules synthesis. In this paper, coculture system of two metabolically engineered Escherichia coli populations were employed which comprises upstream module expressing two enzymes converting para-coumaric acid into resveratrol and the downstream module expressing glucosyltransferase to convert the resveratrol into its glucosidated forms; polydatin and resveratroloside. Upon optimization of the initial inoculum ratio of two E. coli populations, 92 mg resveratrol glucosides/L (236 µM) was produced i.e. achieving 84% bioconversion from 280 µM of p-coumaric acid in 60 h by 3 L fed batch fermentor. This is the report of applying coculture system to produce resveratrol glucosides by expressing the aglycone formation pathway and sugar dependent pathway into two different cells.     

In silico-designed lignin peroxidase from <Emphasis Type="Italic">Phanerochaete chrysosporium</Emphasis> shows enhanced acid stability for depolymerization of lignin

Background

The lignin peroxidase isozyme H8 from the white-rot fungus Phanerochaete chrysosporium (LiPH8) demonstrates a high redox potential and can efficiently catalyze the oxidation of veratryl alcohol, as well as the degradation of recalcitrant lignin. However, native LiPH8 is unstable under acidic pH conditions. This characteristic is a barrier to lignin depolymerization, as repolymerization of phenolic products occurs simultaneously at neutral pH. Because repolymerization of phenolics is repressed at acidic pH, a highly acid-stable LiPH8 could accelerate the selective depolymerization of recalcitrant lignin.

Results

The engineered LiPH8 was in silico designed through the structural superimposition of surface-active site-harboring LiPH8 from Phanerochaete chrysosporium and acid-stable manganese peroxidase isozyme 6 (MnP6) from Ceriporiopsis subvermispora. Effective salt bridges were probed by molecular dynamics simulation and changes to Gibbs free energy following mutagenesis were predicted, suggesting promising variants with higher stability under extremely acidic conditions. The rationally designed variant, A55R/N156E-H239E, demonstrated a 12.5-fold increased half-life under extremely acidic conditions, 9.9-fold increased catalytic efficiency toward veratryl alcohol, and a 7.8-fold enhanced lignin model dimer conversion efficiency compared to those of native LiPH8. Furthermore, the two constructed salt bridges in the variant A55R/N156E-H239E were experimentally confirmed to be identical to the intentionally designed LiPH8 variant using X-ray crystallography (PDB ID: 6A6Q).

Conclusion

Introduction of strong ionic salt bridges based on computational design resulted in a LiPH8 variant with markedly improved stability, as well as higher activity under acidic pH conditions. Thus, LiPH8, showing high acid stability, will be a crucial player in biomass valorization using selective depolymerization of lignin.

     

Determination of creatinine and other uremic toxins in human blood sera with micellar electrokinetic capillary electrophoresis

We have been interested in the clinical use of capillary electrophoresis (CE) to monitor low-molecular-mass uremic toxins in body fluids. Creatinine, an important clinical marker for renal failure, is zwitterionic over a fairly wide pH range (pH 5–9) and can not be resolved from neutral components using free solution CE under these conditions. We report here a micellar electrokinetic capillary chromatography method using an sodium dodecyl sulfate-borate buffer system at pH 9.0 to determine creatinine levels in human serum. This method, performed on deproteinized sera, is also suitable for determining multiple ionic components. Moreover, this method compares favorably with an enzymatic method for creatinine performed in a clinical laboratory and thus appears to be a promising method in terms of potential clinical use.     

Improving microalgae removal efficiency using chemically-processed clays

Chemically-processed clays (PRCs) by base and acid treatments were developed to effectively remove microalgae, and their removal efficiencies were evaluated and compared to that of natural clay. The processed clays were produced by using sodium hydroxide and different amounts of sulfuric acid, and their characteristics were analyzed. Microalgae removal efficiency of the clay was varied depending on the amount of sulfuric acid used in treatments. When adding 1 g/L of a PRC-D type (treated with 1.5 v/w sulfuric acid) to the Microcystis aeruginosa suspension (1 × 10⁶ cells/mL), the highest removal performance (87.3 ± 1.5%) without significant pH variation of the suspension was found among tested PRCs, whereas that of natural clay was 32.1 ± 2.2%. Finally, when selected PRC-D showing best efficiency was applied to lake water taken at a local algae bloomed lake, it exhibited 77.6% removal efficiency of microalgae, indicating 1.7 times higher than the natural clay. The results suggest that processed clays in this study could contribute to effective removal of microalgae in the algal-bloomed area with minimal environmental impact.     

South Asia as a Reservoir for the Global Spread of Ciprofloxacin-Resistant Shigella sonnei: A Cross-Sectional Study

BackgroundAntimicrobial resistance is a major issue in the Shigellae, particularly as a specific multidrug-resistant (MDR) lineage of Shigella sonnei (lineage III) is becoming globally dominant. Ciprofloxacin is a recommended treatment for Shigella infections. However, ciprofloxacin-resistant S. sonnei are being increasingly isolated in Asia and sporadically reported on other continents. We hypothesized that Asia is a primary hub for the recent international spread of ciprofloxacin-resistant S. sonnei.ConclusionsThis study suggests that a single clone, which is widespread in South Asia, is likely driving the current intercontinental surge of ciprofloxacin-resistant S. sonnei and is capable of establishing endemic transmission in new locations. Despite being limited in geographical scope, our work has major implications for understanding the international transfer of antimicrobial-resistant pathogens, with S. sonnei acting as a tractable model for studying how antimicrobial-resistant Gram-negative bacteria spread globally.     

Establishment of a Wolbachia Superinfection in Aedes aegypti Mosquitoes as a Potential Approach for Future Resistance Management

Wolbachia pipientis is an endosymbiotic bacterium estimated to chronically infect between 40–75% of all arthropod species. Aedes aegypti, the principle mosquito vector of dengue virus (DENV), is not a natural host of Wolbachia. The transinfection of Wolbachia strains such as wAlbB, wMel and wMelPop-CLA into Ae. aegypti has been shown to significantly reduce the vector competence of this mosquito for a range of human pathogens in the laboratory. This has led to wMel-transinfected Ae. aegypti currently being released in five countries to evaluate its effectiveness to control dengue disease in human populations. Here we describe the generation of a superinfected Ae. aegypti mosquito line simultaneously infected with two avirulent Wolbachia strains, wMel and wAlbB. The line carries a high overall Wolbachia density and tissue localisation of the individual strains is very similar to each respective single infected parental line. The superinfected line induces unidirectional cytoplasmic incompatibility (CI) when crossed to each single infected parental line, suggesting that the superinfection would have the capacity to replace either of the single constituent infections already present in a mosquito population. No significant differences in fitness parameters were observed between the superinfected line and the parental lines under the experimental conditions tested. Finally, the superinfected line blocks DENV replication more efficiently than the single wMel strain when challenged with blood meals from viremic dengue patients. These results suggest that the deployment of superinfections could be used to replace single infections and may represent an effective strategy to help manage potential resistance by DENV to field deployments of single infected strains.     

Apolipoprotein A-II Plus Lipid Emulsion Enhance Cell Growth via SR-B1 and Target Pancreatic Cancer In Vitro and In Vivo

Background

Apolipoprotein A-II (ApoA-II) is down regulated in the sera of pancreatic ductal adenocarcinoma (PDAC) patients, which may be due to increase utilization of high density lipoprotein (HDL) lipid by pancreatic cancer tissue. This study examined the influence of exogenous ApoA-II on lipid uptake and cell growth in pancreatic cancer (PC) both in vitro and in vivo.

Methods

Cryo transmission electron microscopy (TEM) examined ApoA-II’s influence on morphology of SMOFLipid emulsion. The influence of ApoA-II on proliferation of cancer cell lines was determined by incubating them with lipid+/-ApoA-II and anti-SR-B1 antibody. Lipid was labeled with the fluorophore, DiD, to trace lipid uptake by cancer cells in vitro by confocal microscopy and in vivo in PDAC patient derived xenograft tumours (PDXT) by fluorescence imaging. Scavenger receptor class B type-1(SR-B1) expression in PDAC cell lines and in PDAC PDXT was measured by western blotting and immunohistochemistry, respectively.

Results

ApoA-II spontaneously converted lipid emulsion into very small unilamellar rHDL like vesicles (rHDL/A-II) and enhanced lipid uptake in PANC-1, CFPAC-1 and primary tumour cells as shown by confocal microscopy. SR-B1 expression was 13.2, 10.6, 3.1 and 2.3 fold higher in PANC-1, MIAPaCa-2, CFPAC-1 and BxPC3 cell lines than the normal pancreatic cell line (HPDE6) and 3.7 fold greater in PDAC tissue than in normal pancreas. ApoA-II plus lipid significantly increased the uptake of labeled lipid and promoted cell growth in PANC-1, MIAPaCa-2, CFPAC-1 and BxPC3 cells which was inhibited by anti SR-B1 antibody. Further, ApoA-II increased the uptake of lipid in xenografts by 3.4 fold.

Conclusion

Our data suggest that ApoA-II enhance targeting potential of lipid in pancreatic cancer which may have imaging and drug delivery potentialities.