Immunocytochemical detection of the growth-associated protein B-50 by newly characterized monoclonal antibodies in human brain and muscle

The growth-associated protein B-50 also termed GAP-43, F1, pp46, P-57 and neuromodulin is a nervous tissuespecific protein kinase C (PKC) substrate that is considered to play a major role in neurite formation, regeneration, and neuroplasticity. We describe the isolation of seven mouse monoclonal antibodies (Mabs) directed against B-50. The Mabs are produced against the bovine B-50, selected by ELISA for cross-reactivity with its human counterpart, and evaluated on Western blots in comparison with the well-characterized affinity-purified rabbit polyclonal antibodies to rat-B-50. The Western blots show that the Mabs NM1, NM4, and NM6 recognize specifically the B-50 of bovine, human, and rat brain extract and the purified PKC phosphorylated and unphosphorylated rat B-50 isoforms. The Mabs NM2 and NM3 cross-react with bovine B-50 immunoreactive c-kinase substrate (BICKS), a protein sharing a 17 amino acid sequence homology with B-50. Two Mabs are useful for the detection of B-50 immunoreactivity in formalin-fixed human and rat brain tissues. In human specimen of the hippocampus, a characteristic neuropil distribution of B-50 is detected by the Mabs. In human muscle, Mabs reveal B-50 in nerve bundles and in axons at motor end plates. Thus, these Mabs are useful in investigating the function and localization of the B-50 protein.     

Non-random distribution of individual genetic diversity along an environmental gradient

Improving our knowledge of the links between ecology and evolution is especially critical in the actual context of global rapid environmental changes. A critical step in that direction is to quantify how variation in ecological factors linked to habitat modifications might shape observed levels of genetic variability in wild populations. Still, little is known on the factors affecting levels and distribution of genetic diversity at the individual level, despite its vital underlying role in evolutionary processes. In this study, we assessed the effects of habitat quality on population structure and individual genetic diversity of tree swallows (Tachycineta bicolor) breeding along a gradient of agricultural intensification in southern Québec, Canada. Using a landscape genetics approach, we found that individual genetic diversity was greater in poorer quality habitats. This counter-intuitive result was partly explained by the settlement patterns of tree swallows across the landscape. Individuals of higher genetic diversity arrived earlier on their breeding grounds and settled in the first available habitats, which correspond to intensive cultures. Our results highlight the importance of investigating the effects of environmental variability on individual genetic diversity, and of integrating information on landscape structure when conducting such studies.     

Failure of testicular development associated with a rearrangement of 9p24.1 proximal to the SNF2 gene

In 46,XY individuals, testes are determined by the activity of the SRY gene (sex-determining region Y), located on the short arm of the Ychromosome. The other genetic components of the cascade that leads to testis formation are unknown and may be located on the Xchromosome or on the autosomes. Evidence for the existence of several loci associated with failure of male sexual development is indicated by reports of 46,XY gonadal dysgenesis associated with structural abnormalities of the Xchromosome or of autosomes (chromosomes9, 10, 11 and 17). In this report, we describe the investigation of a child presenting with multiple congenital abnormalities, mental retardation and partial testicular failure. The patient had a homogeneous de novo 46,XY,inv dup(9)(pter→p24.1::p21.1 →p23.3::p24.1→qter) chromosome complement. No deletion was found by either cytogenetic or molecular analysis. The SRY gene and DSS region showed no abnormalities. Southern blotting dosage analysis with 9p probes and fluorescent in situ hybridisation data indicated that the distal breakpoint of the duplicated fragment was located at 9p24.1, proximal to the SNF2 gene. We therefore suggest that a gene involved in normal testicular development and/or maintenance is present at this position on chromosome 9. Received: 20 January 1997 / Accepted: 5 November 1997     

Photoaffinity spin-labeling of the Ca2+ ATPase in sarcoplasmic reticulum : Evidence for oligomeric structure

A spin labeled fatty acid (16-doxylstearic acid) was linked to a photochemical reacting group (azido derivative). When the molecule is introduced, at a low concentration, into rabbit sarcoplasmic reticulum membranes, the spectrum before illumination is identical to the spectrum obtained with the corresponding spin labeled fatty acid. After illumination, a large immobilized components is seen. It corresponds to about 70% of the ESR signal of the effectively bound label, at room temperature. The fraction of immobilized component varies with temperature, from 100% at 0°C to 50% at 35°C. Addition of a small amount of detergent (dodecyl octaethylene glycol monoether), under non solubilizing conditions, decreases the fraction of signal due to a strongly immobilized probe. A possible interpretation is that the immobilized signal reflects protein bound spin labels trapped in Ca²⁺ ATPase oligomers, which are partially dissociated by detergent addition or temperature increase.     

Carbohydrate-induced hypertriglyceridemia: modifying factors and implications for cardiovascular risk.

High-carbohydrate/low-fat, isocaloric diets have repeatedly been shown to increase plasma triglyceride concentrations. The present review addresses recent developments relevant to several important unresolved issues. These include the type of dietary carbohydrate that is most likely to induce hypertriglyceridemia, predictors of individual susceptibility, modifiable physiologic factors that may mitigate the plasma triglyceride response, underlying metabolic mechanisms that are responsible for increased plasma triglycerides, and implications of altered serum lipid profiles for atherogenic risk. Although some progress has been made in this field, the central public health issue - the net effect on cardiovascular risk - remains unresolved.     

The prickly pear (Opuntia ficus-indica [Cactaceae]) in South Africa: Utilization of the naturalized weed,and of the cultivated plants

In spite of successful biological control efforts, vast areas of South Africa remain infested with the spiny prickly pear weed, Opuntia ficus-indica. These remnant populations, which vary from sparse to dense and which are more or less stable, are confined mainly to the eastern Cape. They provide a popular fruit and a limited source of income for certain sectors of the population in these areas. In terms of existing legislation, there are, however, restrictions on the large-scale utilization of these fruit. A novel method for mass-rearing the cochineal insect, Dactylopius coccus, for the commercial production of a red dye (carminic acid) has been developed. The cultivated spineless prickly pear has recently been enjoying renewed attention from researchers and growers alike. Whereas it has traditionally been cultivated mainly as a droughtresistant fodder crop, and is still popular as such, it is now increasingly recognised as a fruit, in its own right, with considerable promise as a commercial crop for local and export markets. There is also a need to encourage the use of very young cladodes (nopalitos) as a highly nutritious vegetable for human consumption in South Africa.     

Biotransformation of 2,2-Dimethyl-1,3-Propanediol to 3-Hydroxypivalic Acid by Acetobacter Acetii DSMZ3508 and Related Bacteria

Acetobacter acetii DSMZ3508 and related bacteria converted 2,2-dimethyl-1,3-propanediol into 3-hydroxypivalic acid (2,2-dimethyl-3-hydroxypropionic acid; 3HP) during submerged cultivation in mineral salt medium. The maximum yield of 3-hydroxypivalic acid was 24.4% of the fed substrate after 18 days. Cultivation parameters, as pH, cell density, optimal substrate concentration, and oxygen supply for the bioconversion process were determined.     

Liquefaction of dulse (Palmaria palmata (L.) Kuntze) by a commercial enzyme preparation and a purified endo ,β-1,4-D-xylanase

Liquefaction of dry and freshPalmaria palmata by food grade enzyme preparations and a purified endo--1,4-D-xylanase was studied.The endo--1,4-D-xylanase (EC 3.2.1.8) was purified to homogeneity from a commercial food grade enzyme prepared fromAspergillus niger. It has a molecular weight of 22 500, a pI of 3.5, is inactive toward corn arabinoxylan,p-nitrophenyl--D-xylose, carboxymethyl cellulose but shows a weak activity toward microcrystalline cellulose. It hydrolyzes oat and dulse xylan equally well in seawater and deionized water essentially into xylose and xylobiose. It is stable between pH 5.5 to 9.0 and 0 to 30 °C and its activity is optimal at pH 4.5–5.5 and 40–60 °C. It has a K_m of 2.2 and 2.8 mg ml^-1 and V_max of 3600 and 3900 nkat mg^-1 of protein on oat and dulse xylan, respectively.Acetate buffer, deionized water and seawater alone extracted 62.6 to 64.5 % of the dry weight of dry dulse, but the use of commercial food grade enzyme preparations or the purified xylanase improved liquefaction to 81.2–87.1 %. Xylose and galactose were the only sugars present in the soluble extracts. Deionized and seawater extracted 58.8–52.7 and 39.1–42.2% of the dry weight of the fresh algae collected in fall and summer, respectively. Only galactose was found in the seawater extract, while some xylose with galactose were measured in the deionized water extract of the fresh autumn algal sample. Purified and crude xylanase improved liquefaction of fresh algae to 79.8–81.4 and 71.9–77.9% of the fresh dry weight (fall and summer, respectively) in deionized and seawater, respectively, and increased the xylose content of the soluble fractions. Polysaccharides in the soluble residues were composed of 1,3/1,4-linked xylose, 1-linked galactose (floridoside) and 1,4-linked glucose (cellulose) and contained essentially 1,4-linked xylose and 1,4-linked glucose in insoluble fractions obtained after enzymatic treatment.The use of xylanase-containing food grade enzyme preparations improves liquefaction ofPalmaria palmata, particularly from fresh alga. This study indicates that processing such as drying may modify markedly the solubility ofP. palmata cell wall polysaccharides, which would imply the existence of some organization and/or other components in the fresh cell wall that lower xylan solubility in seawater.     

Gastric inhibitory polypeptide receptor in hamster pancreatic beta cells. Direct cross-linking, solubilization and characterization as a glycoprotein

125I-labelled gastric inhibitory polypeptide (125I-GIP) is directly cross-linked to its specific receptor in hamster pancreatic beta cell membranes by using an ultraviolet irradiation procedure. This approach results in the identification of a GIP-protein complex of apparent Mr 64,000. The labelling of this protein species is specific since it is inhibited when incubating the membranes with increasing doses of native GIP (0.1 nM-1 microM) together with 125I-GIP, half-maximal inhibition being elicited by 5 nM peptide. Reduction of the GIP-protein complex by 100 mM dithiothreitol induces a decrease of the electrophoretic mobility of the complex. Alternatively pretreatment of membranes with dithiothreitol (up to 1 M) does not prevent the binding of 125I-GIP to its receptor. When prelabelled membranes are extracted by 0.5% Triton X-100 (v/v) and the extract is layered on a Sephadex G-50 column, a high peak of radioactivity is eluted with the void volume of the column. Treatment of this peak by 10 min ultraviolet irradiation followed by SDS-PAGE leads to identification of a major band of Mr 64,000. When the peak is further layered on Sephacryl S-200 it yields a single peak of radioactivity corresponding to a protein species with a Stokes radius of 3.2 nm and an apparent Mr of 65,000. The solubilized GIP-receptor complex is specifically adsorbed by Sepharose coupled to wheat germ agglutinin and concanavalin A and eluted from these lectins by their respective sugars. In conclusion the GIP receptor in pancreatic beta cells is a protein monomer of apparent Mr 59 000; its structure is maintained by intrachain disulfide bridges, these bonds being, however, not involved in the interaction of GIP with its receptor; the GIP receptor is a glycoprotein containing N-acetylglucosamine, mannose and probably sialic acid in its carbohydrate moiety.