Novel Pathway of Toluene Catabolism in the Trichloroethylene-Degrading Bacterium G4

o-Cresol and 3-methylcatechol were identified as successive transitory intermediates of toluene catabolism by the trichloroethylene-degrading bacterium G4. The absence of a toluene dihydrodiol intermediate or toluene dioxygenase and toluene dihydrodiol dehydrogenase activities suggested that G4 catabolizes toluene by a unique pathway. Formation of a hybrid species of ¹⁸O- and ¹⁶O-labeled 3-methylcatechol from toluene in an atmosphere of ¹⁸O₂ and ¹⁶O₂ established that G4 catabolizes toluene by successive monooxygenations at the ortho and meta positions. Detection of trace amounts of 4-methylcatechol from toluene catabolism suggested that the initial hydroxylation of toluene was not exclusively at the ortho position. Further catabolism of 3-methylcatechol was found to proceed via catechol-2,3-dioxygenase and hydroxymuconic semialdehyde hydrolase activities.     

Contra-IL 2; a suppressor lymphokine that inhibits IL 2 activity

Suppressive activity of culture supernatant of AS-9 (AS-9 CS), a T cell hybridoma line that was derived from fusion of BW5147 thymoma and splenic T cells of anti-lymphocyte serum-treated C3H mice, was analyzed. AS-9 CS inhibited allogeneic cytotoxic T lymphocyte (CTL) generation as well as T cell proliferation to alloantigens and mitogens, but failed to inhibit B cell response to lipopolysaccharide or growth of tumor and fibroblast cells. Although addition of AS-9 CS to the allogeneic sensitization culture as late as on day 2 of incubation resulted in maximal inhibiton of CTL generation, removal of AS-9 CS on day 3 of incubation abolished its inhibitory effect. Addition of purified IL 2 together with AS-9 CS to the allogeneic sensitization cultures only partially abrogated the suppression. Experiments with IL 2-dependent cytotoxic T cell line (CTLL) showed that AS-9 CS suppressed the IL 2-induced proliferation of CTLL. Preincubation of AS-9 CS with CTLL removed its inhibitory effect on CTL generation. These results indicate that AS-9 CS interferes with the mechanism of T cell activation by IL 2. On this basis, AS-9 CS was named contra-IL 2.     

Dopamine- and octopamine-sensitive adenylate cyclase in the brain of adultCulex pipiens mosquitoes

The effects of dopamine and octopamine on adenylate cyclase activity were studied on the head homogenate of adult Culex pipiens mosquitoes in vitro. Both dopamine and octopamine were shown to increase the cyclic AMP content in the homogenate. The antagonist haloperidol blocked the production of cyclic AMP induced from dopamine but had no effect on the production of cyclic AMP induced by octopamine at the concentrations tested. The opiate agonist etorphine was ineffective at reducing cyclic AMP levels induced by either dopamine or octopamine at the concentrations tested.     

Effect of calcium on the endogenous phosphorylation of mouse brain microsomes in vitro

1. The endogenous phosphorylation of mouse brain microsomes was studied using the technique of acrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS). 2. It was found that specific proteins and lipids in brain microsomes were phosphorylated by the terminal phosphate of ATP under appropriate conditions. Six peaks of radioactivity were observed on SDS-polyacrylamide gel electrophoresis of 32Pi-labelled brain microsomes. The peaks were designated as P-I, P-II, P-III, P-IV, P-V, and P-VI. The peaks from P-I to P-V, which consist of phosphoproteins, underwent rapid dephosphorylation. On the other hand, P-VI, which consists of phospholipids, remained unaffected even after the complete hydrolysis of added ATP. 3. With the addition of 100 muM CaCl2 to the assay medium, the phosphorylation of brain microsomal proteins was stimulated; in the regions of P-I, P-II, and P-III, the amounts of 32Pi incorporation were approximately twice the 32Pi incorporation in the absence of Ca2+. On the other hand, 32Pi incorporation into P-VI was unaffected irrespective of the presence or absence of 100 muM CaCl2. In the presence of higher concentrations of Ca2+ (1-10 mM), the phosphorylation of all components was inhibited.     

Sexual Selection in the Three-Spined Stickleback

The purpose of this study was to seek evidence for the influence of sexual selection on both male and female three-spined sticklebacks, a species in which the ♂ alone has parental responsibilities. We studied the intrasexual relationships of ♂♂ and ♀♀ prior to courtship, and the intrasexual and intersexual relationships of both sexes during courtship. Results demonstrated that both ♂♂ and ♀♀ compete intrasexually, and suggested that both sexes are discriminative in mate selection. These results are compatible with Trivers' (1972) model of the role of parental investment in sexual selection.     

Profile of a nonylphenol-degrading microflora and its potential for bioremedial applications

Nonylphenol (NP) is an important intermediate in the production of various commercial and industrial materials, but is also known as a ubiquitous pollutant in urban aquatic environments. We recently studied the NP-degrading activities of microflora in several aquatic environments, and found a notable degrading activity for wastewater from a sewage treatment plant in Tokyo. This result led us to isolate NP-degrading microbes and identify biodegradation products. Using conventional plate culture techniques and molecular biological methods, Pseudomonas and Sphingomonas species, which are known for their degradation activities of many aromatic compounds, have been isolated. But it has also been found that Sphingomonas sp. (S-strain) is necessary and sufficient for the degradation of NP. Although the role of Pseudomonas sp. (P-strain) remains unclear, P-strain seems to provide some co-nutrients for the growth of S-strain. The degradation products were analyzed by GC/MS and NMR. More than 95% of NP was degraded within 10 days and aromatic compounds other than NP were not found, suggesting that the phenolic part of NP was completely degraded. We also examined the potential of S-strain for bioremedial applications. S-strain cells immobilized on chitosan or alginate beads retain their NP-degrading activity in flask-scale experiments. Furthermore, the chitosan-bound cells in a lab-scale bioreactor have been found to be persistent for repeated use, suggesting that S-strain is applicable to the treatment of NP-contaminated wastewater.     

Novel histone modifications in microglia derived from a mouse model of chronic pain

As the resident immune cells in the central nervous system, microglia play an important role in the maintenance of its homeostasis. Dysregulation of microglia has been associated with the development and maintenance of chronic pain. However, the relevant molecular pathways remain poorly defined. In this study, we used a mass spectrometry-based proteomic approach to screen potential changes of histone protein modifications in microglia isolated from the brain of control and cisplatin-induced neuropathic pain adult C57BL/6J male mice. We identified several novel microglial histone modifications associated with pain, including statistically significantly decreased histone H3.1 lysine 27 mono-methylation (H3.1K27me1, 54.8% of control) and H3 lysine 56 tri-methylation (7.5% of control), as well as a trend suggesting increased H3 tyrosine 41 nitration. We further investigated the functional role of H3.1K27me1 and found that treatment of cultured microglial cells for 4 consecutive days with 1–10 μM of NCDM-64, a potent and selective inhibitor of lysine demethylase 7A, an enzyme responsible for the demethylation of H3K27me1, dose-dependently elevated its levels with a greater than a two-fold increase observed at 10 μM compared to vehicle-treated control cells. Moreover, pretreatment of mice with NCDM-64 (10 or 25 mg/kg/day, i.p.) prior to cisplatin treatment prevented the development of neuropathic pain in mice. The identification of specific chromatin marks in microglia associated with chronic pain may yield critical insight into the contribution of microglia to the development and maintenance of pain, and opens new avenues for the development of novel nonopioid therapeutics for the effective management of chronic pain.