Patterns of expression of the JIM4 arabinogalactan-protein epitope in cell cultures and during somatic embryogenesis in Daucus carota L.

Spatiotemporal patterns of expression of the cell-surface arabinogalactan-protein epitope defined by monoclonal antibody JIM4 (J.P. Knox et al., 1989, Development 106, 47–56) have been characterized by indirect immunofluorescence during the process of somatic embryogenesis in Daucus carota L. The JIM 4 epitope (J4e) occurred on cells established in culture from hypocotyl explants which appeared to derive, at least in part, from the epidermal cells of the hypocotyl. Cultures maintained in the presence of 2,4-dichlorophenoxyacetic acid developed proembryogenic masses of which only infrequent cells at the surface expressed J4e. Sub-culture at a low cell density and withdrawl of the synthetic auxin resulted in an increase in J4e expression in most surface cells and most abundantly in surface layers of cells at the future shoot end of developing embryos. The transition to heart-shaped embryos occurred concurrently with the expression of J4e by groups of cells beneath the developing cotyledons, at the junction of the future root and shoot. At this stage, J4e was also expressed by a single well-defined layer of cells at the surface of the embryos. Advancement to the mature torpedo stage was accompanied by the expression of the epitope on cells forming two regions of the future stele and of cells associated with the cotyledonary provascular tissue characteristic of the carrot seedling. At this stage there was substantially less expression of the marker antigen by epidermal cells, although infrequent expression by isolated cells of the epidermis was maintained. The correlation of J4e expression with the development and distinction of plant tissue patterns during somatic embryogenesis indicates a role for plasma-membrane arabinogalactan proteins in these processes.Abbreviations AGP arabinogalactan protein - 2,4-D 2,4-di-chlorophenoxyacetic acid - J4e JIM 4 epitope - PEM proembryogenic mass We thank Andrew Davis for photographic assistance and Roger Pennell for useful discussions.     

European post-glacial forests: compositional changes in response to climatic change

Abstract. Multivariate analysis of an extensive palyno-logical database for Europe has enabled reconstruction of broad-scale vegetation history. Whereas many major features of present vegetation patterns were established early in the Holocene, floristic composition of the forests has changed continuously up to the present day. For example, although ‘mixed deciduous forests’ had reached approximately their present extent in northwest Europe by 8000 B.P., Tilia peaked in abundance in these forests during the middle post glacial, whereas Pinus was most abundant in these forests during the early post-glacial and Fagus increased in abundance only in recent millennia. Pollen-climate response surfaces for major pollen taxa show how their distribution and abundance patterns relate to contemporary climate. Past forest-compositional changes were responses to climatic changes, the nature of which can be inferred from pollen-climate response surfaces. Post-glacial climate changes have been different in magnitude and direction in different regions of Europe. For example, in recent millennia the vegetation changes indicate decreasing summer temperatures in northern Europe but increasing summer temperatures in the Mediterranean region. The way in which vegetation responded to past climatic changes gives insight into the likely response of vegetation to future climate changes induced by the ‘greenhouse effect’.     

A sensitive and rapid luminescence sandwich assay for antibodies to hepatitis-B surface antigen (Anti-HBsAg)

A chemiluminescent assay for hepatitis-B surface antigen is described which used an isoluminol derivative as the label. The assay is precise intra-assay CV, 1.96-2.45%; inter-assay CV, 5.26-8.11% and has a lower detection limit for hepatitis-B surface antigen of 1.3U/I.     

Cellular Stress Causes Accumulation of the Glucose Transporter at the Surface of Cells Independently of their Insulin Sensitivity

The stimulation of glucose transport in response to various types of stress has been studied. There is no relationship between effects of stress-inducing agents on glucose transport and their effects on cellular protein synthesis. Although the effect of stress on glucose transport appears analogous to its stimulation by insulin, cells that are slightly insulin-sensitive in terms of glucose transport (BHK cells) show a similar degree of stimulation as highly insulin-sensitive cells (differentiated 3T3-L1 cells). External labeling of the transporter protein with a photoactivatable derivative of mannose, 2-N-4-(1-azi-2,2,2-trifluoroethyl) benzoyl-1, 3-bis-(D-mannos-4-yloxy)-propylamine, shows that most of the increased glucose transport activity correlates with an increase in the amount of the transporter on the cell surface. Cells subjected to K⁺-depletion, which inhibits endocytosis and results in an accumulation of receptors at the cell surface, show the same increase in glucose transport as cells exposed to stress; stressed cells show no further increase in glucose transport when subjected to K⁺ depletion. These results support the view (Widnell, C.C., Baldwin, S.A., Davies, A., Martin, S., Pasternak, C.A. 1990. FASEB J 4:1634–1637) that cellular stress increases glucose transport by promoting the accumulation of glucose transporter molecules at the cell surface. Received: 20 June 1995/Revised: 29 September 1995     

Attachment of Pasteuria penetrans Endospores to the Surface of Meloidogyne javanica Second-stage Juveniles

Pasteuria penetrans spore adhesion to Meloidogyne javanica second-stage juveniles (J2) was examined following several different pretreatments of the latter. The detergents sodium dodecyl sulfate and Triton X-100, the carbohydrates fucose and α-methyl-D-mannoside, and the lectins concanavalin A and wheat germ agglutinin reduced spore attachment. Spores exposed to M. javanica surface coat (SC) extract exhibited decreased adherence to the J2 surface. Second-stage juveniles that had been treated with antibodies recognizing a 250-kDa antigen of J2 SC extract had fewer spores attached to their surfaces, as compared to nontreated J2, except in the head region. This inhibition pattern was similar to that of antibody-labelling on M. javanica J2 as observed by electron microscopy. It is suggested that several SC components, such as carbohydrate residues, carbohydrate-recognition domains, and a 250-kDa antigen, are involved in P. penetrans spore attachment to the surface of M. javanica.     

Nitrogen losses from perennial grass species

Nitrogen losses from plants may occur through a variety of pathways, but so far, most studies have only quantified losses of nutrients by above-ground litter production. We used ¹⁵N pulse labelling to quantify total nitrogen losses from above- and below-ground plant parts. Using this method we were able to include also pathways other than above-ground litter production. To test the hypothesis that species from nutrient-poor habitats lose less nitrogen than species from more fertile soils, six perennial grasses from habitats with a wide range of nutrient availability were investigated: Lolium perenne, Arrhenatherum elatius, Anthoxanthum odoratum, Festuca rubra, F. ovina and Molinia caerulea. The results of an experiment carried out in pots in a green-house at two fertility levels show that statistically significant losses occur through pathways other than above-ground litter production. In the low fertility treatment, most (70%) losses from L. perenne occurred by litter production, but in Ar. elatius, F. rubra, F. ovina and M. caerulea, more than 50% of labelled N losses took place by root turn-over, leaching or exudation from roots. When nutrient supply increased, the ¹⁵N losses in above-ground dead material increased in all species and in Ar. elatius, A. odoratum and F. rubra the ¹⁵N losses via other pathways decreased. Ranked according to decreasing turnover coefficient the sequence of species was: L. perenne, A. odoratum, F. rubra, F. ovina, Ar. elatius, M. caerulea. These results suggest that species adapted to sites with low availability of nutrients lose less nitrogen (including above- and below-ground losses) than species adapted to more fertile soils.     

Adaptive surface antigen variation in Mycoplasma bovis to the host immune response

Abstract The variability of predominant Mycoplasma bovis surface antigens in the presence of specific immune pressure was analyzed in an in vitro assay to determine if M. bovis could escape immune destruction. We have shown that serum antibodies from immunized or experimentally infected calves and monoclonal antibodies which specifically react with previously characterized or as yet undefined major M. bovis membrane surface proteins cause repression of expression or shortening of the target protein, or induce switching to expression of an antigenically distinct variant protein. We have further demonstrated that removal of the inducing antibody results in reversion to the original phenotype. These results suggest that the level of expression and the length of M. bovis surface antigens in the host is modulated by cognate antibodies. According to the surface antigenic variation systems, random selection of preexisting variants resistant to antibody-mediated inhibition or direct regulation of gene expression may be means by which this organism evades host immune defences.     

Comparison of a Bone Criterion and a dental Criterion for estimation of age at death in the study of an Ancient Cemetery

Two methods, a visual method based on the appearance of the auricular surface of the ilium and a metric method based on two dental criteria, were used in conjunction for estimation of skeletal age at death in paleodemographic study of an ancient cemetery, and were found to be coherent. However, the paleodemographic profiles differed according to sex, indicating a sexual difference in the evolution of the sacroiliac joint.     

Interstitial noradrenaline concentration of rat hearts as influenced by cellular catecholamine uptake mechanisms

Marked concentration differences of noradrenaline (NA) between the vascular and the interstitial compartment were detected by sampling interstitial transudate from isolated perfused rat hearts. The ratios of vascular/interstitial concentration amounted to 7.4 to 1.3 depending on the concentration of NA administered (3 × 10^–9 to 10^–6 M). These concentration differences were abolished by inhibitors of uptake₁ desipramine (DMI) I and uptake, (O-methyl-isoprenaline (OMI)). Neuronal uptake, was characterized by a K_m of 0.22 mol/l and a V_max of 370 pmol × min^–1 × gWWT^–1, extraneuronal uptake₂ by a K_UPTAKE of = 0.313 min^–4.The apparent permeability surface area (P×S)-product calculated from uptake rate and transcapillary concentration difference was significantly decreased by administrating 100 mol/l (NA) in presence of DMI. A presumed endothelial uptake mechanism contributing to catecholamine translocation was investigated in endothelial cells in culture. These cells showed a specific noradrenaline uptake with a K_m of 4.35 mol/l and a V_max of about 75 pmol × min^–1 x gWWT^–1. Any inhibiton by inhibitors of both of the two noradrenaline uptakes was lacking. The uptake rate of this mechanism is insufficient to contribute to the diffusive conductivity of the capillary wall (P × S-product). We conclude from our investigations on interstitial concentrations of catecholamines and transcapillary concentration differences, that the capillary wall, owing to its metabolic and diffusional characteristics, influences the exchange of catecholamines to a substantial and physiologically relevant extent.     

Photoaffinity labelling of a cytokinin-binding integral membrane protein in plant mitochondria

Two target polypeptides were detected by photoaffinity labelling of purified mung bean mitochondria using tritiated 2-azido-N⁶-benzylaminopurine. SDS-PAGE and fluorography of total mitochondrial proteins after the photoaffinity reaction showed a labelled 32 kDa polypeptide (intensely labelled) and a 57 kDa polypeptide (less intensely labelled). The latter was assumed to be the and/or subunit of F₁ATPase since it was the most abundant polpeptide in gels stained with Coomassie Blue. Partial purification of F₁ATPase demonstrated that the 32 kDa polypeptide was not a component of the ATPase complex. Fractionation experiments showed that the 32 kDa protein was integrally associated with mitochondrial membranes and could be enriched by simple washing and detergent extraction procedures.