Modulation of adenylate cyclase/cyclic AMP response by thyrotropin and prostaglandin E2 in cultured thyroid cells. 1. Negative regulation.

Isolated porcine thyroid cells, cultured in the presence of thyrotropin (greater than or equal to 0.25 mU/ml) or prostaglandin E2 (greater than or equal to 0.1 micron), showed decreased adenosine 3':5'-monophosphate (cyclic AMP) response to further thyrotropin or prostaglandin E2 stimulation, respectively. Kinetics of the refractory process to thyrotropin and prostaglandin E2 are different: (a) maximal refractoriness to prostaglandin E2 was attained after 2--6 h exposure to prostaglandin E2 while refractoriness to thyrotropin was maximal only after 12--24 h; (b) the degree of refractoriness to prostaglandin E2 was much greater than that to thyrotropin. Refractoriness to thyrotropin or prostaglandin E2 is characterized: by specificity for each thyroid stimulator; by dependence upon the dose of thyrotropin or prostaglandin E2 in culture, e.g. induction of high degree of refractoriness with 0.5 mU/ml thyrotropin (or 1 micron prostaglandin E2), which elicits only a small cyclic AMP increase; by time requirement for induction; by partial effect; by changes of maximum activation of cyclic AMP response; by reversibility. This refractoriness of the cyclic AMP response was not induced by dibutyryl adenosine 3':5'-monophosphate. It was not attributed to increased cyclic AMP-phosphodiesterase activity, but to alterations in the receptor-adenylate cyclase system. Prevention of refractoriness to thyrotropin or prostaglandin E2 by incubation of cells in the presence of actinomycin D, puromycin and cycloheximide suggests that new RNA and protein syntheses are required for the development of the refractory state.     

Respiration-dependent uncoupler-stimulated ATPase activity in castor bean endosperm mitochondria and submitochondrial particles

1. The uncoupler-stimulated ATPase activity of castor bean endosperm mitochondria and submitochondrial particles has been studied. The rate of ATP hydrolysis catalyzed by intact mitochondria was slow and little enhanced by addition of uncouplers at the concentration required for uncoupling the oxidative phosphorylation. ATPase activity was stimulated at higher concentrations of uncouplers.

2. 1-Anilinonaphthalene 8-sulfonate fluorescence was decreased when the mitochondria were oxidizing succinate. Carbonylcyanide-p-trifluoromethoxyphenylhydrazone and antimycin reversed the succinate-induced fluorescence diminution. ATP did not induce the fluorescence response.

3. The addition of succinate, NADH or ascorbate/N,N,N′,N′-tetramethyl-p-phenylenediamine as electron donor induced high ATPase activity in the presence of low concentrations of uncouplers. Stimulating effect of uncouplers was completely abolished by further addition of antimycin.

4. Submitochondrial particles were prepared by sonication. The particles catalyzed a rapid hydrolysis of ATP and carbonylcyanide-p-trifluoromethoxyphenylhydrazone at 10^-8 M did not stimulate the ATPase activity. Addition of succinate induced uncoupler-stimulated ATPase activity. The effect of succinate was completely abolished by further addition of antimycin.

5. The treatment of submitochondrial particles by trypsin or high pH also induced uncoupler-stimulated ATPase activity.

6. The above results were interpreted to indicate that ATPase inhibitor regulated the back-flow reaction of mitochondrial oxidative phosphorylation.     

Myeloperoxidase has Directly-opposed Effects on Nitration Reaction--Study on Myeloperoxidase-deficient Patient and Myeloperoxidase-knockout Mice

Myeloperoxidase (MPO) catalyzes a nitration reaction to form nitrotyrosine in the presence of high nitrite, the metabolite of NO. Human leukocyte was shown to cause phenolic nitration using released MPO as a catalyst in the presence of nitrite. It opposes our previous finding that inhibition of MPO was essential for phenol nitration in human leukocyte study. To clarify the role of MPO, we utilized MPO-deficient human leukocytes and MPO-knockout mice. Even in the absence of exogenously added nitrite, high nitration product was observed in MPO-deficient leukocytes. In liver subjected to ischemia/reperfusion injury, a significantly higher amount of nitrotyrosine was produced in MPO-knockout mice than in normal mice. These results clearly demonstrate that MPO inhibits the accumulation of nitration products in vivo . Further experiments showed that MPO could degrade nitrotyrosine in the presence of glutathione. Thus, MPO-induced degradation of nitration products may cause the underestimation of the nitration product generated in vivo . We conclude that MPO may act predominantly to scavenge nitrotyrosine under physiological nitrite condition, and protect against injurious effect of nitrotyrosine.     

Synthesis of Methylenecyclobutyl- and Cyclobutenyl Adenine,Potent Antiviral Carbocyclic Analogues of Oxetanocin A

Abstract

9-Cyclobutyladenines bearing both methylene and hydroxymethyl groups, 3 and 4, were prepared by dehydration of carbocyclic oxetanocin A (1a). Introduction of a double bond into cyclobutane ring was achieved by allylic oxidation of N ⁶-benzoyl-9-[3-methylenecyclobutyl]adenine (12), which after several steps, afforded 9-[3-(hydroxy-methyl)-2-cyclobutenyl)adenine (5).     

Estimation of carbon biomass and community structure of planktonic bacteria in Lake Biwa using respiratory quinone analysis

The relationship between bacterial respiratory quinone (RQ) concentration and biomass was assessed for Lake Biwa bacterial assemblages to evaluate the utility of bacterial RQ concentration as an indicator of bacterial carbon. The biomass estimated from the RQ concentration correlated well with that from cell volume, indicating that RQ concentration is an appropriate indicator of bacterial biomass. The estimated carbon content per unit of RQ (carbon conversion factor) of bacteria was 0.67 mg C nmol RQ^?1. Bacterial carbon biomass, which was estimated from the RQ concentration using the conversion factor, ranged between 0.008 and 0.054 mg C L^?1 (average 0.025 mg C L^?1) at 5 m depth and between 0.010 and 0.024 mg C L^?1 (average 0.015 mg C L^?1) at 70 m depth. Ubiquinone-8-containing bacteria dominated the epilimnion and hypolimnion. Compared to conventional image analysis, bacterial RQ analysis is a less laborious method of simultaneously determining bacterial biomass and community.     

Differentially Expressed Proteins Associated with Fusarium Head Blight Resistance in Wheat

Background

Fusarium head blight (FHB), mainly caused by Fusarium graminearum, substantially reduces wheat grain yield and quality worldwide. Proteins play important roles in defense against the fungal infection. This study characterized differentially expressed proteins between near-isogenic lines (NILs) contrasting in alleles of Fhb1, a major FHB resistance gene in wheat, to identify proteins underlining FHB resistance of Fhb1.

Methods

The two-dimensional protein profiles were compared between the Fusarium-inoculated spikes of the two NILs collected 72 h after inoculation. The protein profiles of mock- and Fusarium-inoculated Fhb1⁺NIL were also compared to identify pathogen-responsive proteins.

Results

Eight proteins were either induced or upregulated in inoculated Fhb1⁺NIL when compared with mock-inoculated Fhb1⁺NIL; nine proteins were either induced or upregulated in the Fusarium-inoculated Fhb1⁺NIL when compared with Fusarium-inoculated Fhb1⁻NIL. Proteins that were differentially expressed in the Fhb1⁺NIL, not in the Fhb1⁻NIL, after Fusarium inoculation included wheat proteins for defending fungal penetration, photosynthesis, energy metabolism, and detoxification.

Conclusions

Coordinated expression of the identified proteins resulted in FHB resistance in Fhb1⁺NIL. The results provide insight into the pathway of Fhb1-mediated FHB resistance.     

Dynamical Properties of Condensed Charged Polymer Melts

Abstract

Dynamical properties of condensed charged polymer melts are studied with a two-dimensional model and molecular dynamics simulation. Screened Coulombic interactions are assumed for the interactions between the monomer charges as well as the counterions which were introduced to neutralize the total monomer charges of polymer chains. Through molecular dynamics calculations, we have obtained the radial distribution function and velocity auto-correlation function, and their density dependences. As a structural characteristics in condensed charged polymer melts, we find that the monomers tend to form triangular structures locally. The radial distribution function for the center monomers implies that the polymer chains on our two-dimensional model are not entangled. The diffusion properties for both the counter-ions and innermost monomers are studied in detail. In this paper, we have also computed neutral polymer melts to study the effects of the presence of the long-range Coulombic interactions on the dynamical properties of polymer melts. We find that the Coulombic interactions significantly reduce the self diffusion. Snapshots analyses obtained from the molecular dynamics simulation suggest that the reptation model is not a proper model for two-dimensional polymer-chain melts, in which there are little entanglement effects in agreement with the result of the analyses for the radial distribution function.