Oleosin expression and trafficking during oil body biogenesis in tobacco leaf cells

We have established a versatile method for studying the interaction of the oleosin gene product with oil bodies during oil body biogenesis in plants. Our approach has been to transiently express a green fluorescent protein (GFP)-tagged Arabidopsis oleosin gene fusion in tobacco leaf cells containing bona fide oil bodies and then to monitor oleosin-GFP expression using real-time confocal laser scanning microscopy. We show that normally non-oil-storing tobacco leaf cells are able to synthesize and then transport oleosin-GFP fusion protein to leaf oil bodies. Synthesis and transport of oleosin-GFP fusion protein to oil bodies occurred within the first 6 h posttransformation. Oleosin-GFP fusion protein exclusively associated with the endoplasmic reticulum and was trafficked in a Golgi-independent manner at speeds approaching 0.5 microm sec(-1) along highly dynamic endoplasmic reticulum positioned over essentially static polygonal cortical endoplasmic reticulum. Our data indicate that oil body biogenesis can occur outside of the embryo and that oleosin-GFP can be used to monitor early events in oil body biogenesis in real-time.     

Dietary fibre on cell proliferation in large bowel mucosal crypts near or away from lymphoid nodules and on mineral bioavailability

The effect of consumption for 24 weeks of different amounts (0%, 5% or 10% w/w) of fermentable (pectin and guar gum) or nonfermentable (cellulose and lignin) dietary fibres on cell proliferation and other parameters in large bowel mucosal crypts was studied in rats. In all 12 dietary groups, the crypts located over the distal aggregate of lymphoid nodules (ALN) had more colchicine arrested metaphase figures per midaxial crypt section (MC) and a longer crypt column height than crypts located three to four cm away from this ALN. These differences are attributed to the tropic influence of nodular cells in the ALN. Consumption of fermentable fibre decreased pH in the lumen of the caecum, and glucose, Zn and Cu in serum but increased Ca and Mg in serum. The decrease in caecal pH and serum glucose was significantly correlated with a decrease in MC. Increased intake of the nonfermentable fibre types increased faecal bulk but had no significant correlation with the other measured crypt parameters. Multiple regression analyses was used to model the relationships between the mucosal crypt criterion variables and the two measured predictor variables, caecal pH and serum glucose. Relationships between dietary fibre, ALN, MC, bioavailability of dietary minerals and risk of colorectal cancer are discussed.     

The biology and use of the African brush-tailed porcupine (Atherurus africanus, Gray, 1842)as a food animal. A review

The brush-tailed porcupine (Atherurus africanus) is a hystricomorph rodent, which frequents the forests of West and Central Africa. With an average weight of 3 kg, it is a favourite source of meat for urban and rural populations of Gabon, Nigeria, Cameroon or Congo. Hunted in large quantities, its price is often higher than that of other game or domestic animals. Although its current productivity in captivity is limited to a single young per birth and two to three births per year per female, this species could be a good candidate for minilivestock programmes in African forest areas if its current reproductive potential in captivity could be improved. Further research should be encouraged on its biology and reproduction since the current level of hunting for this species is probably not sustainable. Captive breeding programmes could play a role in assessing a better knowledge of the species' biology, and in reducing the effects of intensive hunting in areas where this activi ty is no longer sustainable.     

Mechanisms involved in the photosensitized inactivation of Acanthamoeba palestinensis trophozoites

Aims:  To advance our understanding of the mechanisms involved in the RLP068 phthalocyanine-photosensitized inactivation of Acanthamoeba palestinensis trophozoites through a precise identification of the targets of the photoprocess in both the cytosolic and mitochondrial compartments.
Methods and Results:  We followed the activities of selected marker enzymes as well as we performed fluorescence and transmission electron microscopy investigations of the alterations induced by the photoprocess in the fine structure of subcellular compartments. RLP068 is preferentially located in the contractile vacuole: the fluorescence in that site is particularly evident in the unirradiated cells and becomes more diffused after irradiation. Electron microscopic analysis of photosensitized A. palestinensis cells clearly shows that the swelling of trophozoites and the appearance of vacuoles spread throughout the cytoplasm after phototreatment. The activity of a typical cytoplasmic enzyme, such as lactate dehydrogenase, underwent a 35% decrease as a consequence of the photoprocess, reflecting the photodamage induced by migrating phthalocyanine molecules in their micro-environment.
Conclusions:  The presence of multiple targets for the phthalocyanine-photosensitized process is of utmost importance because this pattern of cell damage makes it unlikely that photoresistant A. palestinensis strains are gradually selected or mutagenic phenomena are developed as a consequence of the photoinduced damage.
Significance and Impact of the Study:  Photosensitization via phthalocyanines appears to represent an efficient and safe approach for achieving a close control of the population of a potentially pathogenic protozoan such as A. palestinensis , opening new perspectives for the disinfection of microbiologically polluted waters.     

2,4-Diaminopyrimidine MK2 inhibitors. Part I: Observation of an unexpected inhibitor binding mode

MK2 is a Ser/Thr kinase of significant interest as an anti-inflammatory drug discovery target. Here we describe the development of in vitro tools for the identification and characterization of MK2 inhibitors, including validation of inhibitor interactions with the crystallography construct and determination of the unique binding mode of 2,4-diaminopyrimidine inhibitors in the MK2 active site. Use of these tools in the optimization of a potent and selective inhibitor lead series is described in the accompanying Letter.     

On temporal partitioning of a community of ectomycorrhizal fungi

Several mechanisms may contribute to the high species richness often reported in ectomycorrhizal (ECM) fungal communities, including spatial and temporal partitioning. Here, we focus on temporal partitioning. Using molecular methods, we determined the frequencies of occurrence of ECM fungal species detected as hyphae and ECM roots in the forest floor of a Pinus resinosa plantation during a 13-month period. We then used a novel statistical procedure to place the most frequently occurring ECM fungal species into groups distinguished by their patterns of relative frequency over time. Three groups with contrasting temporal patterns were distinguishable for fungal species detected as hyphae. Two groups were distinguishable for species detected as ECM roots. Our results support the hypothesis that temporal partitioning occurs among the species of ECM fungi in this community, but we did not address its causes, which may have involved interactions among species' physiological tolerances, temporal environmental variability, temporal patterns of root production, and variation in fungal genet lifespan. These interactions should be the subjects of future research.     

Contrasting below-ground views of an ectomycorrhizal fungal community

Ectomycorrhizal fungal communities have been characterized in a number of ways. Here we compare colonized root-tip and mycelia views of an ectomycorrhizal fungal community. Ectomycorrhizal fungi, both as mycelia and colonized root tips, were identified in soil samples taken from a pine plantation. We determined that for some ectomycorrhizal fungal species multiple root tips from a single soil sample were not independent. Therefore in the comparison of root-tip and mycelia views, we considered species to be present or absent from each soil sample irrespective of the number of root tips colonized by the species. We observed 39 ectomycorrhizal fungal species in total, but 12 were observed exclusively as mycelia and 11 exclusively colonizing root tips. The relative frequencies of 10 species occurring as both mycelia and root tips were not independent of the method of observation. Our results suggest that ectomycorrhizal fungal species differ in their spatial distributions on root tips, and that root-tip and mycelia views of the community are different.     

Model structures of alpha-2 adrenoceptors in complex with automatically docked antagonist ligands raise the possibility of interactions dissimilar from agonist ligands

Antagonist binding to alpha-2 adrenoceptors (alpha2-ARs) is not well understood. Structural models were constructed for the three human alpha2-AR subtypes based on the bovine rhodopsin X-ray structure. Twelve antagonist ligands (including covalently binding phenoxybenzamine) were automatically docked to the models. A hallmark of agonist binding is the electrostatic interaction between a positive charge on the agonist and the negatively charged side chain of D3.32. For antagonist binding, ion-pair formation would require deviations of the models from the rhodopsin structural template, e.g., a rotation of TM3 to relocate D3.32 more centrally within the binding cavity, and/or creation of new space near TM2/TM7 such that antagonists would be shifted away from TM5. Thus, except for the quinazolines, antagonist ligands automatically docked to the model structures did not form ion-pairs with D3.32. This binding mode represents a valid alternative, whereby the positive charge on the antagonists is stabilized by cation-pi interactions with aromatic residues (e.g., F6.51) and antagonists interact with D3.32 via carboxylate-aromatic interactions. This binding mode is in good agreement with maps derived from a molecular interaction library that predicts favorable atomic contacts; similar interaction environments are seen for unrelated proteins in complex with ligands sharing similarities with the alpha2-AR antagonists.