Phytanic acid alpha-oxidation and complementation analysis of classical Refsum and peroxisomal disorders

Summary We have measured the production of ¹⁴CO₂ from exogenous [1-¹⁴C] phytanic acid in fibroblast monolayers from patients with classical Refsum's disease and peroxisomal disorders. Activities in the different disorders were (percentage of control): classical Refsum's disease (5%), isolated peroxisomal acyl-CoA oxidase deficiency (75%), Zellweger syndrome (4%), neonatal adrenoleukodystrophy (5%), and rhizomelic chondrodysplasia punctate (3%). Absence of complementation was demonstrated between Zellweger syndrome and infantile Refsum's disease lines after polyethylene glycol fusion, with decreases of average activity of 11% relative to unfused cell mixtures. Classical Refsum's disease, rhizomelic chondrodysplasia punctata, and neonatal adrenoleukodystrophy lines all complemented one another, and Zellweger syndrome or infantile Refsum's disease lines, with average activity increases of 522%–772%. No intragenic complementation was observed within either group. Four complementation groups were detected suggesting that at least four genes are involved in phytanic acid -oxidation: one gene for the enzyme phytanic acid -hydroxylase (probably mitochondrial); one gene for a regulatory factor for the expression of phytanic acid -decarboxylation activity and two membrane-bound peroxisomal enzymes involved in the synthesis of plasmalogens; two genes for the assembly of functional peroxisomes and/or import of proteins into peroxisomes.     

c-fos expression in the amygdala:In vivo antisense modulation and role in anxiety

Summary 1. The amygdaloid complex is a key structure in mechanisms of fear and anxiety. Expression of the immediate-early gene c-fos has been reported in the central nucleus of the amygdala following various stressors, but the functional role of this phenomenon has remained unknown.2. c-fos expression was observed in the central nucleus when rats were subjected to a pharmacologically validated animal model of anxiety, the Vogel conflict test, but not after mere exposure to the test apparatus. Bilateral amygdala injection of a 15-mer phosphorothioate c-fos antisense oligodeoxynucleotide prior to testing blocked conflict-induced c-fos expression and had behavioral effects similar to those of established antianxiety drugs.3. Separate experiments determined that antisense treatment did not affect conflict behavior by acting on shock thresholds or drinking motivation.4. These findings provide evidence that neuronal activation and c-fos induction in the amygdala may be of importance for mechanisms of fear and anxiety.     

Population regulation of a tropical damselfly in the larval stage by food limitation,cannibalism, intraguild predation and habitat drying

The relative importance of intraspecific, interspecific, and seasonal causes of larval mortality were investigated for aquatic larvae of the giant damselfly Megaloprepus coerulatus in Panama. These larvae live in water-filled holes in fallen and living trees, where they and three other common odonate species are the top predators. By mid wet season, M. coerulatus larvae were found in nearly half of all tree holes that harbored odonates. Although M. coerulatus were typically, but not always, eliminated from holes inhabited by larger hetero-specifics, M. coerulatus were more likely to encounter conspecifics than other odonate species. Hole with less than 11 of water rarely contained more than a single larva. In large holes where M. coerulatus was the only odonate species present, multiple larvae coexisted at a density of one larva per 1–21 of water. There the absence of 2–4 of the 5 larval size classes, despite a continuous input of eggs, suggested that cannibalism was a common cause of mortality. The size of the final instar, which determined adult body size, was correlated positively with tree hole volume for male, but not female, larvae. Experiments showed that when two larvae were placed together in 0.4–1 holes with abundant tadpole prey, the larger larva killed the smaller one. Often the larva that was killed was not eaten. Small larvae were more tolerant of each other than were pairs of medium or large larvae. Before killing occurred, the presence of larger larvae reduced the growth of smaller individuals, relative to controls. Obligate killing was density-dependent. In 3.0–1 holes with ad libitum prey, conspecific killing occurred until the larval density stabilized at one larva per 1–1.5 I, similar to the density found in large holes under field conditions, For M. coerulatus, cannibalism functions to reduce the number of potential competitors for food in addition to providing nutrition. When interactions between paired larvae in small holes were experimentally prevented, competition for food reduced the growth of one or both larvae relative to controls. Holes that were watered during the dry season supported larval densities similar to those in the wet season. Thus, dry season mortality could not be attributed to a decrease in available prey. Rather, M. coerulatus larvae could not survive more than 1 month of complete drying. Because the dry season typically lasts more than 6 weeks, habitat drying is a secondary source of mortality, affecting second- or third-generation larvae that fail to emerge before tree holes dry out completely.     

Human familial and sporadic breast cancer: analysis of the coding regions of the 17β-hydroxysteroid dehydrogenase 2 gene (EDH17B2) using a single-strand conformation polymorphism assay

17-Hydroxysteroid dehydrogenase (17HSD) is one of the key enzymes in estrogen metabolism, catalyzing the reversible reaction between estradiol and the less active estrogen, estrone. The gene encoding this enzyme, EDH17B2, has been mapped to chromosome 17, region q12–q21, in the vicinity of BRCA1, an as yet unidentified gene that appears to be involved in familial breast cancer and in familial ovarian cancer. The possibility that EDH17B2 gene is the same as BRCA1 was tested by screening for mutations in the coding regions of EDH17B2, using a polymerase chain reaction/single-strand conformation polymorphism method. An AG transition creating a new BstUI site at exon 6 was the only frequent sequence alteration found in the coding region of the gene. This mutation also led to an amino acid substitution of serine to glycine at position 312 (312^S312^G) in the 17HSD protein. Since the nucleotide change was detected both in specimens from patients with familial or sporadic cancer and in control samples, and at similar rates, this mutation appears to be of a polymorphic nature. In addition, a rare polymorphism located at intron 5 was detected. This CT substitution creates a BbvI site and is not thought to have any effect on 17HSD activity. The results indicate that there are no major alterations in the coding areas of EDH17B2 and thus studies testing the hypothesis that EDH17B2 may be the same as BRCA1 should be extended to the promoter and regulatory elements of EDH17B2.     

Nuclear and chloroplast transformation inChlamydomonas reinhardtii: strategies for genetic manipulation and gene expression

Transformation of the nuclear, chloroplast, and mitochondrial genomes can now be accomplished inChlamydomonas reinhardtii. Many biosynthetic pathways are carried out in the chloroplast, and efforts to manipulate these pathways will require that gene products be directed to this compartment. Chloroplast proteins are encoded in either the chloroplast or nuclear genome. In the latter case they are synthesized in the cytoplasm and imported post-translationally into the chloroplast. Thus, strategies for expressing foreign genes or overexpressing endogenous genes whose products reside in the chloroplast could involve either genome. This paper reviews the present status of transformation methodology for the nuclear and chloroplast genomes inChlamydomonas. Considerations for expressing gene products in the chloroplast are discussed. Experimental evidence for homologous recombination during transformation of the nuclear genome is presented.     

Morphological phylogenetics of theAsteraceae-Asteroideae,with notes on character evolution

The relationships within theAsteraceae-Asteroideae are scrutinized by means of cladistic analyses of morphological and chemical data. New data are presented, and novel interpretations of features previously used in cladistic analyses are made. Examples are insertion of anther filaments in the corolla, and morphology of disc floret styles, respectively. The results include a polyphyletic or paraphyleticInuleae s. l., and a paraphyleticHeliantheae s.l. in which theEupatorieae constantly form a subclade. Comparisons are made with previous cladistic analyses of morphological data, and those acquired from molecular data. Speculations about the evolution of certain organs are presented.     

Feeding by the pine weevil Hylobius abietis in relation to sun exposure and distance to forest edges

Abstract 1 The intensity of feeding by adult pine weevils Hylobius abietis (L.) on the stem bark of Norway spruce Picea abies (L.) Karst. seedlings planted in rows with a north–south orientation across a clear‐cutting, was measured throughout a growth season. The feeding was then correlated to light interception, soil temperature and distance to the nearest forest edge. 2 Feeding was at least twice as intense on seedlings in the central part of the clear‐cutting compared to those at the edges. The decline began approximatety 15 m from the edge and was of similar proportions on both the sun‐exposed and shaded sides. 3 Measures of global radiation and soil temperature correlated well with consumption on the shaded side. However, on the sun‐exposed side, there were no apparent correlations with global radiation or soil temperature that could explain the decline in consumed bark area. 4 We conclude that the decline in feeding towards the forest edges was mainly due to factors other than the microclimate variables we monitored. We suggest that the presence of roots of living trees along the forest edge may reduce damage to seedlings, since they provide an alternative source of food for the weevils. This alternative‐food hypothesis may also explain why seedlings in shelterwoods usually suffer less damage from pine weevils than seedlings in clear‐cuttings.     

Psednotrichia, a genus of the tribe Senecioneae hitherto misplaced in the Astereae (Asteraceae)

Anderberg, A. A. & Karis, P. O. 1995. Psednotrichia , a genus of the tribe Senecioneae hitherto misplaced in the Astereae (Asteraceae). — Nord. J. Bot. 15: 375–379. Copenhagen. ISSN 0107–055X.
The tribal position for the little known genus Psednorrichia , is established. This monotypic genus, hitherto placed in the tribe Astereae, is a congener of Xyridopsis of the tribe Senecioneae (Asteraceae). The two species of Xyridopsis are here transferred to Psednotrichia and the new combinations P. xyridopsis and P. newtonii are made. A brief discussion of the morphology of the genus, and its systematic position within the tribe Senecioneae is provided.     

Three-dimensional structure prediction of the NAD binding site of proton-pumping transhydrogenase from Escherichia coli

A three-dimensional structure of the NAD site of Escerichia coli transhydrogenase has been predicted. The model is based on analysis of conserved residues among the transhydrogenases from five different sources, homologies with enzymes using NAD as cofactors or substrates, hydrophilicity profiles, and secondary structure predictions. The present model supports the hypothesis that there is one binding site, located relatively close to the N-terminus of the α-subunit. The proposed structure spans residues α145 to α287, and it includes five β-strands and five α-helices oriented in a typical open twisted α/β conformation. The amino acid sequence following the GXGXXG dinucleotide binding consensus sequence (residues α172 to α177) correlates exactly to a typical fingerprint region for ADP binding βαβ folds in dinucleotide binding enzymes. In the model, aspartic acid α195 forms hydrogen bonds to one or both hydroxyl groups on the adenosine ribose sugar moiety. Threonine α196 and alanine α256, located at the end of βB and βD, respectively, create a hydrophobic sandwich with the adenine part of NAD buried inside. The nicotinamide part is located in a hydrophobic cleft between αA and βE. Mutagenesis work has been carried out in order to test the predicted model and to determine whether residues within this domain are important for proton pumping directly. All data support the predicted structure, and no residue crucial for proton pumping Was detected. Since no three-dimensional structure of transhydrogenase has been solved, a well based tertiary structure prediction is of great value for further experimental design in trying to elucidate the mechanism of the energy-linked proton pump. © 1995 Wiley-Liss, Inc.     

Isolation and partial characterization of bovine gingival AB collagen

Summary Limited proteolysis with pepsin solubilized 25% of the insoluble gingival matrix as mainly soluble collagenous material. Fractional salt precipication at neutral pH resulted in the separation of types III and I at 1.8 and 2.6 M NaCl, respectively. In addition, a collagenous fraction accounting for 2% of the solubilized collagen and precipitating at 4.5 M NaCl was shown to be identical with type V collagen. Isolation and partial characterization of the constituent-α-chains of the 4.5 M PPT by gel filtration, ion exchange and hydroxylapatite chromatography as well as disc electrophoresis showed that gingival type V collagen contains αA and αB chains in a ratio αB/αA of 1.73–1.8. Electron microscopic examination of ATP-precipitates showed that this collagen type gave only one kind of SLS aggregates with asymmetric band pattern characteristically different from that of type I collagen. The data provide evidence that gingival AB collagen is a heteropolymer in which the αA and αB chains are assembled in the same macromolecule in a 1∶2 ratio.