Survival and Nodulating Ability of Indigenous and Inoculated Rhizobium leguminosarum biovar trifolii in Sterilized and Unsterilized Soil Treated with Sewage Sludge

Rhizobium leguminosarum biovar trifolii was detected in soil from 41 of 47 plots, within nine sewage sludge-treated sites with different soil characteristics and heavy metal contents. However, although population size varied widely, there was no consistent correlation with soil heavy metal concentration. Indigenous populations in 20 plots within four selected sites retained their ability to induce effective nodule formation after incubation of soil in the dark for 165 days. In sterilized (γ-irradiated) soil, Rhizobium survival varied from 0.01% to 95% depending on the soil sample and strain used. Metal-resistant strains with non-mucoid colonies survived less well than mucoid metal-sensitive strains. Received: 26 June 2000/Accepted: 31 July 2000     

3-(7-Azaindolyl)-4-arylmaleimides as potent, selective inhibitors of glycogen synthase kinase-3

A novel series of acyclic 3-(7-azaindolyl)-4-(aryl/heteroaryl)maleimides was synthesized and evaluated for activity against GSK-3beta and selectivity versus PKC-betaII, as well as a broad panel of protein kinases. Compounds 14 and 17c potently inhibited GSK-3beta (IC(50)=7 and 26 nM, respectively) and exhibited excellent selectivity over PKC-betaII (325 and >385-fold, respectively). Compound 17c was also highly selective against 68 other protein kinases. In a cell-based functional assay, both 14 and 17c effectively increased glycogen synthase activity by inhibiting GSK-3beta.     

Habituation of western gorillas: the process and factors that influence it

Habituation of western gorillas to human presence is generally an expensive, lengthy and difficult process. Here we describe the habituation process for two groups of western gorillas at the Mondika Research Center, with the hope that the lessons we learned will facilitate future gorilla studies. We expand upon earlier studies by describing the process through complete habituation for both males and females, and for more than one group. The major obstacle to habituation was developing sufficient tracking skills to follow gorilla trail on a daily basis. Once this was achieved, the silverback became semi-habituated (i.e. ignoring human presence during half of contacts) within a year, although the majority of group females continued to avoid humans. As female presence at contacts increased, a period of male recidivism followed, requiring an additional year before his complete habituation was reached. Habituating the females took longer than the male, but we found, contrary to earlier studies, that it consisted of the same stages, including avoidance, aggression, and curiosity before habituation. We compare results between groups and across sites and discuss how factors such as tracking abilities, group size and cohesion, population density and home range overlap, and the manner of approaching gorillas during contacts influence the habituation process.     

Identification of the GPR55 agonist binding site using a novel set of high-potency GPR55 selective ligands

Marijuana is the most widely abused illegal drug, and its spectrum of effects suggests that several receptors are responsible for the activity. Two cannabinoid receptor subtypes, CB1 and CB2, have been identified, but the complex pharmacological properties of exogenous cannabinoids and endocannabinoids are not fully explained by their signaling. The orphan receptor GPR55 binds a subset of CB1 and CB2 ligands and has been proposed as a cannabinoid receptor. This designation, however, is controversial as a result of recent studies in which lysophosphatidylinositol (LPI) was identified as a GPR55 agonist. Defining a biological role for GPR55 requires GPR55 selective ligands that have been unavailable. From a β-arrestin, high-throughput, high-content screen of 300000 compounds run in collaboration with the Molecular Libraries Probe Production Centers Network initiative (PubChem AID1965), we identified potent GPR55 selective agonists. By modeling of the GPR55 activated state, we compared the GPR55 binding conformations of three of the novel agonists obtained from the screen, CID1792197, CID1172084, and CID2440433 (PubChem Compound IDs), with that of LPI. Our modeling indicates the molecular shapes and electrostatic potential distributions of these agonists mimic those of LPI; the GPR55 binding site accommodates ligands that have inverted-L or T shapes with long, thin profiles that can fit vertically deep in the receptor binding pocket while their broad head regions occupy a horizontal binding pocket near the GPR55 extracellular loops. Our results will allow the optimization and design of second-generation GPR55 ligands and provide a means for distinguishing GPR55 selective ligands from those interacting with cannabinoid receptors.     

Molecular Mechanism by Which a Potent Hepatitis C Virus NS3-NS4A Protease Inhibitor Overcomes Emergence of Resistance

Although optimizing the resistance profile of an inhibitor can be challenging, it is potentially important for improving the long term effectiveness of antiviral therapy. This work describes our rational approach toward the identification of a macrocyclic acylsulfonamide that is a potent inhibitor of the NS3-NS4A proteases of all hepatitis C virus genotypes and of a panel of genotype 1-resistant variants. The enhanced potency of this compound versus variants D168V and R155K facilitated x-ray determination of the inhibitor-variant complexes. In turn, these structural studies revealed a complex molecular basis of resistance and rationalized how such compounds are able to circumvent these mechanisms.     

Physiological changes in oocyte-cumulus cell complexes from diabetic mice that potentially influence meiotic regulation

We have previously shown that the type I diabetic condition significantly alters meiotic regulation in mouse oocytes. In the present study, possible physiological deficiencies underlying such meiotic dysfunction were examined in oocyte-cumulus cell complexes (OCC) from type I diabetic mice. Whereas the diabetic condition did not affect glycolysis or the tricarboxylic acid cycle, the increased flux of glucose through the pentose phosphate pathway in response to FSH treatment was suppressed. De novo purine synthesis was also compromised, and ATP levels were reduced in freshly isolated OCC. Additionally, diabetes resulted in a reduction in FSH-mediated cAMP synthesis. The responsiveness of the oocyte to cAMP was also affected; fewer oocytes were induced to resume maturation after a stimulatory pulse with cAMP analogs. Meiotic induction triggered by FSH was significantly reduced, but that stimulated by phorbol ester or epidermal growth factor was affected to a much lesser extent. In addition to metabolic deficiencies, the cell-cell communication between the oocyte and the cumulus cells was reduced in diabetic mice as determined by coupling assays. Thus, numerous physiological parameters are affected by type I diabetes, and these changes may collectively contribute to altered meiotic regulation.