Thyroid hormone regulation of Na,K-ATPase subunit-mRNA expression in neonatal rat myocardium

Summary Regulation of Na,K-ATPase mRNA isoform and mRNA expression by thyroid hormone (T₃) in neonatal rat myocardium was examined. In euthyroid neonates between ages of 2 and 5 days, mRNA₁, mRNA₃, and mRNA₁ abundances were nearly constant while mRNA₂ was undetectable. During the interval between postnatal days 5 and 15, mRNA₃ decreased to negligible levels and mRNA₂ became expressed and increased in abundance to account for 20% of the mRNA pool by the 15^th postnatal day. To examine the effect of T₃ on this developmental program, neonates were injected with 75 g T₃/100 g body weight or diluent alone on the second and third postnatal days and myocardial Na,K-ATPase subunit-mRNA abundances were determined on the third and fourth postnatal days. Because T₃ treatment increased the RNA/DNA ratios of myocardial tissue, the subunit-mRNA abundances were normalized per unit DNA. Following 24 and 48 hr of T₃ treatment, the abundances of mRNA₁, mRNA₃, and mRNA₁ increased, while mRNA₂ continued to remain undetectable during the 2-day interval between the second to fourth postnatal days. It is concluded that T₃ augments the abundance of Na,K-ATPase subunit mRNAs that are already being expressed in the neonatal rat myocardium. The results further suggest that T₃ does not act as a molecular switch in the developmental expression of the mRNA isoforms in rat myocardium during the first four postnatal days.     

Hyalocyten in der Gewebekultur

Zusammenfassung Hyalocyten wurden aus Glaskörper und Retina von Hühner- und Kaninchenembryonen sowie von Kälbern gezüchtet. Sie entwickelten sich in vitro über sog. Ausgangszellschichten der Retina von jungen zu reifen Formen. Diese Zellen gehören zur Gruppe der Makrophagen.Eine Dauerzüchtung scheint nur in Verbindung mit den Ausgangszellschichten möglich, da bei reifen Hyalocyten keine Zellteilungen mehr zu beobachten sind.

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Hyalocytes in tissue culture

Summary Hyalocytes from the vitreous body and retina of chick and rabbit embryos as well as of calves have been cultured. They develop in vitro from immature to mature forms on primary cell layers of the retina. These elements belong to the macrophage group.Permanent culture seems to be only successful in connection with the primary cell layer since cell division is no longer observed in mature ldhyalocytes.

Herrn Prof. Dr. Ernst Lindner und Herrn Dr. Wollensack danke ich für die Anregung zu dieser Arbeit, Frl. Edith Leschinski, Herrn Manfred Meisinger und Frau Anneliese Rehberger für ihre ausgezeichnete Mitarbeit bei der Durchführung der Untersuchungen.     

Micropropagation of mature Chinese tallow tree (Sapium sebiferum Roxb.)

An in vitro propagation technique based on axillary bud proliferation has been developed for matureSapium sebiferum trees. Nodal segments cultured on Murashige and Skoog (MS) medium supplemented with benzyl adenine (1–10 m and -naphthaleneacetic acid (0–0.5 m showed axillary bud proliferation. Shoots proliferated in vitro were multiplied on Murashige and Skoog medium containing 2.5 m benzyl adenine and 0.25 m -naphthaleneacetic acid. Seasonal changes affected the shoot proliferation potential of the initial explant. Shoots were rooted on a half-strength, growth-regulator-free, agar-gelled, MS medium after a 48-h treatment on half-strength MS liquid medium with 10 m indole-3-butyric acid. Rooted plantlets were potted and acclimatized in a growth chamber and then moved to the greenhouse. Four-month-old plants were transplanted to the field.Abbreviations BA Benzyl adenine - IBA Indole-3-butyric acid - 2-ip N⁶-(-dimethylallylamino)purine - MS Murashige and Skoog (1962) medium - NAA -Naphthaleneacetic acid     

Immunochemical comparison of membrane-associated and secreted arabinogalactan-proteins in rice and carrot

Arabinogalactan-proteins (AGPs) occurring in suspension-cultured rice (Oryza saliva L.) cells, their conditioned medium and at the rice root apex were investigated using monoclonal antibodies and the AGP-binding -glucosyl Yariv reagent ( GlcY). A monoclonal antibody, LM2, was generated that recognized an acidic carbohydrate epitope common to two soluble AGPs occurring in the conditioned medium of proliferating rice cells, membrane-associated AGPs (rmAGP) in the cultured cells and two AGPs at the rice root apex. In addition, LM2 recognized AGPs secreted by suspensioncultured carrot (Daucus carota L.) cells. The two AGPs of the rice culture medium, srAGP1 and srAGP2, were discriminated by their mobilities during sodium dodecyl sulfate-polyacrylamide gel electrophoresis, reaction with GlcY, the presence of arabinogalactan epitopes and anion-exchange chromatography. The association of rmAGP with the plasma membrane was investigated by Triton-X-114/aqueous partitioning of both microsomal and plasma-membrane preparations and rmAGP was found to partition into the detergent phase, indicating that AGPs are hydrophobic plasma-membrane proteins in rice. This was in contrast to plasma-membrane AGPs of suspension-cultured carrot cells that partitioned into the aqueous phase. At the rice root apex most of the AGP was associated with the microsomal fraction and also partitioned into the detergent phase, although a distinct highmolecular-mass AGP entered the aqueous phase.Abbreviations AGP arabinogalactan-protein - GlcY -glucosyl Yariv reagent - ELISA enzyme-linked immunosorbent assay We gratefully acknowledge support from the Leverhulme Trust, the UK Biotechnology and Biological Sciences Research Council and the Royal Society.     

α-Subunit Composition of Nicotinic Cholinoreceptors of Neurons of the Guinea Pig Inferior Mesenteric Ganglion: an Immunohistochemical Study

Using immunoperoxidase labeling, we studied the subunit composition of nicotinic acetylcholine receptors, nAChR, in preparations of the inferior mesenteric ganglion, IMG, of the guinea pig. Antibodies against synthetic peptides corresponding to agonist-binding membrane components of the ₃, ₄, ₅, and ₇ nAChR subunits were used. The presence of ₃-specific antibodies was revealed on the membranes of about 58% of large neurons and of all small ganglionic cells (means of the greater and smaller diameters of the somata 53.8 ± 1.8 vs 33.6 ± 1.4 m, n = 20, and 14.1 ± 0.5 vs 7.5 ± 0.4 m, n = 50, respectively). Labeled cells of the rostral node were distributed evenly, while those of the caudal node were localized mostly within the regions of branching of the lumbar, colonic, and both hypogastric tracts. Immune labels to the ₄ subunit were observed only on the membranes of small ganglionic cells distributed mostly in the region of the internodal commissural tracts. ₅-Specific labeling was found on the membranes of about 63% large neurons, whose distribution was similar to that of the ₃-labeled units, and on all small cells. Immunoreactivity to the ₇ subunit was observed only on the membranes of small cells concentrated around unlabeled large neurons in the region of branching of the intermesenteric, colonic, and both hypogastric tracts. Thus, nAChR in the guinea pig IMG include ₃, ₄, ₅, and ₇ subunits. The nAChR with ₃ and ₅ subunits are localized on the membranes of large ganglionic neurons, whose number and topographical distribution are very close to each other. Our data agree with our results of earlier electrophysiological experiments and are indicative of the crucial role of the ₃- and ₅-containing nAChR in synaptic transmission via the ganglion under study. The presence of the ₄- and ₇-containing nAChR was found only on small ganglionic cells (which are, probably, not the relay units) and their processes.     

Novel cross-strand three-purine stack of the highly conserved 5′-GA/AAG-5′ internal loop at the 3′-end termini of Parvovirus Genomes

We have used two-dimensional nuclear magnetic resonance (2D-NMR), distance geometry (DG) and molecular dynamics / energy minimization (MD/EM) methods to study a 2×3 asymmetric internal loop structure of the highly conserved `5-(GA)/(AAG)-5 bubble' present at the 3-end hairpin of the single-stranded DNA genome of parvoviruses. This motif contains an unpaired adenosine stacked between two bracketed sheared GA pairs. However, the phenomenal cross-strand G-G and A-A stacking in the tandem sheared GA pairs has undergone considerable change. A novel three-purine stacking pattern is observed instead; the inserted A18 base is completely un-stacked from its neighboring G17 and A19 bases, but well stacked with the cross-strand A4 and G3 bases to form a novel A4/A18/G3 stack that is different from the double G/G, A/A or quadruple G/G/G/G stack present in the 5-(GA)/(AG)-5 or 5-(GGA)/(AGG)-5 motifs. Unlike the bulged purine residue that usually causes about 20 degree kink in the helical axis of the parent helix when bracketed by canonical GC or AT base pairs, no significant kink is observed in the present helix containing a bulged-adenine that is bracketed by sheared G A pairs. The phosphodiesters connecting G3-A4 and G17-A18 residues adopt unusual torsional angles close to the trans domain, yet that connecting A18-A19 residues resumes the normal (g <sup>–</sup>) value. The well structured `5-(GAA)/(AG)-5' internal loop in the parvovirus genomes explains its resistance to single-strand specific endonuclease susceptibility.     

A cDNA encoding vacuolar type β-d-fructofuranosidase (Osβfruct3) of rice and its expression in Pichia pastoris

A vacuolar type -d-fructofuranosidase (Osfruct3) was cloned from etiolated rice seedlings cDNA library. It encodes an open reading frame of 688 residues. The deduced amino acid sequence had 58% identity to the vacuolar type -d-fructofuranosidase of maize (Ivr1). Osfruct3 exists as a single copy per genome. Northern analyses showed that Osfruct3 undergoes organ-specific expression and is involved in the adjustment of plant responses to environmental signals and metabolizable sugars. Osfruct3 was also heterologously expressed in Pichia pastoris. The recombinant proteins were confirmed to be a vacuolar type -d-fructofuranosidase.     

Ultrastructural localization of corticotropin, β-lipotropin,and αand β-endorphin in the porcine anterior pituitary

Summary The peroxidase-antiperoxidase immunocytochemical technique was used to identify the ACTH/endorphin cells in the porcine pituitary at the ultrastructural level and to determine the precise subcellular localization of the pro-ACTH/endorphin fragments. The cells display different aspects: 1) large, regular shapes with numerous and large secretory granules; 2) small, irregular and angular shapes with small granules aligned along the periphery of the cell; and 3) intermediate forms. The presence of and -endorphin not only in the same cells but also in the same secretory granules that contain ACTH and -LPH clearly indicates that both the precursor or its fragments and the abovementioned peptides are stored in the same granules and released simultaneously by the corticotropic cells. The presence of FSH in some corticotropic cells is also discussed.Abbreviations used in this Article ACTH corticotropin - -MSH -melanotropin (ACTH I–I3) - CLIP corticotropin-like intermediate lobe peptide (ACTH 18–39) - -LPH -lipotropin - -MSH -melanotropin (-LPH 41–58); -endorphin (-LPH 61–91); -endorphin (-LPH 61–76)     

Chlorophyll mutations in mungbean (Vigna radiata (L.) Wilczek)

Summary Twenty mutants isolated from Latisail, Jhingasail and Pankaj varieties of rice (Oryza sativa L.) were screened for two aspects of nutritive quality, namely crude protein content and distribution pattern of protein in the endosperm. Observations revealed a wide variation for both characters, and while there was no consistent association between protein content and test grain weight, which varied between varieties, a positive correlation between protein content and grain sterility was noted. In a few mutants protein distribution was observed to be varied and showed a similarity to optimum milling characteristics.     

A comparison of two ATPase based schemes for histochemical muscle fibre typing in various mammals

Summary A comparative study was performed on the fibre populations in tibialis anterior muscles of mouse, rat, guinea pig, rabbit, cat and dog using the two different methods of histochemical staining for myofibrillar ATPase after acid (Brooke and Kaiser 1970) or alkaline preincubations (Guth and Samaha 1970). For all species a complete correspondence existed between type I (Brooke and Kaiser 1970) and fibres (Samaha et al. 1970). Gross correspondence (>85%) existed between IIA and IIB (Brooke and Kaiser 1970) and and fibres (Samaha et al. 1970) respectively in mouse, guinea pig, rabbit, cat and dog. In the case of mouse and dog, this high degree of correspondence was based on the assumption that mouse tibialis anterior contains no type I and the dog no type IIB fibres. For the rat, a pronounced overlap existed between IIA fibres on the one hand and and fibres on the other hand as well as between IIB fibres and and fibres. These observations lead to the conclusion that the two classification schemes are not interchangeable for all species and that the two terminologies should be used only in relation with the methods from which they were derived.     

CHO cell growth and recombinant interferon-γ production: Effects of BSA, Pluronic and lipids

The role of bovine serum albumin in mammalian cell cultures and the possibility of its substitution by other components in a serum-free medium has been investigated. In this study, BSA was shown to be important for growth and product formation in CHO cells expressing recombinant human interferon-. There were indications that its stimulating growth effect was dependent on the source of BSA used and probably was related to the purification procedure used for the production of the desired albumin fraction. Cell growth did not occur in the absence of BSA but at low concentration (1 mg ml^–1) it was stimulated by the addition of a combination of a commercial lipid mixture plus Pluronic F68. However, under the latter conditions IFN- production was adversely effected. The importance of individual lipid components was investigated using a statistical approach based on a Plackett-Burman design. Linoleic acid was identified as a positive variable for cell growth while cholesterol was identified as a negative variable for both cell growth and IFN- production. When a combination of linoleic acid plus Pluronic F68 was included in the formulation of low BSA medium, cell growth was similar to that at high BSA concentration (5 mg ml^–1) but the IFN- concentration was significantly reduced (ca. 45%).Abbreviations IFN- interferon- - CHO Chinese Hamster Ovary cells - BSA bovine serum albumin - FAF-BSA fatty acid-free bovine serum albumin     

Consequences of harvesting for genetic diversity in American ginseng (<Emphasis Type="Italic">Panax quinquefolius</Emphasis> L.): a simulation study

American ginseng, Panax quinquefolius L., is one of the most heavily traded medicinal plants in North America. The effect of harvest on genetic diversity in ginseng was measured with a single generation culling simulation program. Culling scenarios included random harvest at varying levels, legal limit random harvest and legal limit mature plant harvest. The legal limit was determined by the proportion of legally harvestable plants per population (% mature plants per population). Random harvest at varying levels resulted in significant loss of genetic diversity, especially allelic richness. Relative to initial levels, average within-population genetic diversity (H _e) was significantly lower when plants were culled randomly at the legal limit (Mann–Whitney U=430, p<0.001) or when only mature plants were culled (Mann–Whitney U=394, p<0.01). Within-population genetic diversity was significantly higher with legal limit mature plant harvest (H _e=0.068) than when plants were culled randomly at the legal limit (H _e=0.064; U=202, p<0.01). Based on these simulations of harvest over one generation, we recommend that harvesting fewer than the proportion of mature plants could reduce the negative genetic effects of harvest on ginseng populations.     

Phytochrome-mediated induction of ascorbate oxidase in different organs of a dicotyledonous seedling (Sinapis alba L.)

Summary The time courses of the level of ascorbate oxidase (AO; EC 1.10.3.3.) were followed in the different organs (cotyledons, hypocotyl, taproot) of the developing mustard seedling. Phytochrome (operationally, far-red light, cf. [20]) rapidly and strongly enhances the rate of apparent ascorbate oxidase (AO)¹ synthesis in cotyledons and hypocotyl, while in the taproot the detectable amount of AO is only small. However, the relative increase of AO as mediated by continuous far-red light is the same in all organs. Far-red dark kinetics indicate that the phytochrome-induced enzyme is much less stable in the hypocotyl than in the cotyledons, at least during the experimental period. It is concluded that the effect of phytochrome on enzyme induction is precisely the same in cotyledons and hypocotyl, while the processes of enzyme degradation are specific for the organ. Thus the time courses of enzyme levels can be determined by the nature of the particular organ, even if no isoenzymes are involved and the mechanism of the inductive process is the same in the different organs.     

Zinc accumulation in the telencephalon of lizards

Summary The zinc concentration in the brains of two species of lizard was determined by atomic-absorption spectrophotometry. The zinc concentration was found to be highest in the telencephalon of Lacerta galloti (21.1 g/g fresh weight) and Podarcis hispanica (16.77±0.8 g/g) while the mesencephalon and brain stem exhibited lower zinc concentrations, i.e., 7.0 g/g in Lacerta galloti and 6.08±0.4 g/g in Podarcis hispanica. This high telencephalic concentration of zinc is paralleled by intense and well-defined Timm reactivity used for demonstrating the presence of zinc-containing boutons at the light-microscope level. Volumetricdensitometric studies of these Timm-reactive zones were performed using serial transverse sections of the same lizard brains.     

Purification, properties and possible gene assignment of an α1,3-fucosyltransferase expressed in human liver

1,3-Fucosyltransferase solubilized from human liver has been purified 40 000-fold to apparent homogeneity by a multistage process involving cation exchange chromatography on CM-Sephadex, hydrophobic interaction chromatography on Phenyl Sepharose, affinity chromatography on GDP-hexanolamine Sepharose and HPLC gel exclusion chromatography. The final step gave a major protein peak that co-chromatographed with 1,3-fucosyltransferase activity and had a specific activity of 5–6 µmol min^–1 mg^–1 and anM _r 44 000 deduced from SDS-PAGE and HPLC analysis. The purified enzyme readily utilized Gal1-4GlcNAc, NeuAc2-3Gal1-4GlcNAc and Fuc1-2Gal1-4GlcNAc, with a preference for sialylated and fucosylated Type 2 acceptors. Fuc1-2Gal1-4Glc and the Type 1 compound Gal1-3GlcNAc were very poor acceptors and no incorporation was observed with NeuAc2-6Gal1-4GlcNAc. A polyclonal antibody raised against the liver preparation reacted with the homologous enzyme and also with the blood group Lewis gene-associated 1,3/1,4-fucosyltransferase purified from the human A431 epidermoid carcinoma cell line. No cross reactivity was found with 1,3-fucosyltransferase(s) isolated from myeloid cells. Examination by Northern blot analysis of mRNA from normal liver and from the HepG2 cell line, together with a comparison of the specificity pattern of the purified enzyme with that reported for the enzyme expressed in mammalian cells transfected with theFuc-TVI cDNA, suggests a provisional identification ofFuc-TVI as the major 1,3-fucosyltransferase gene expressed in human liver.Died June, 1991     

Style morph frequencies in Minnesota populations of Lythrum (Lythraceae)

Style morph frequencies (shortmidlong) were determined for a total of n = 11 918 plants in 16 Minnesota populations of Lythrum salicaria L. Nine populations were in the establishment phase, with population sizes ranging from n = 56 to n = 2 192. Most of these populations exceeded previously reported population sizes in the native European habitat. A nonparametric statistical test, the chi-square (²), can be used to determine if populations are at isoplethic equilibrium (111, shortmidlong); a ² value >5.99 is significant at the 5% level. Only one established population (White Bear Lake, n = 1991, ² = 3.0) fitted the null hypothesis for isoplethy, although all established populations contained all three style morphs. Pooled values for these populations indicated an excess of mids and longs, with shorts being deficient. Colonizing populations had a higher percentage of mids (54%) when compared to established populations (33.7%). Short styles were almost nonexistent (8%) in colonizing populations. Five out of the seven populations lacked at least one style morph. A review of the literature reporting style morph frequencies in tristylous L. salicaria revealed that no statistical analysis for isoplethy has been performed. Darwin originally assumed that all populations would be isoplethic, possessing equal numbers of all three style morphs, but concluded, without statistical analysis, that, instead, populations were anisoplethic. Since tests for statistical deviations from the expected frequencies (111) have not been used, ² analysis was performed. Several of these populations were at isoplethic equilibrium (Nadder su2 = 1.7, Blelham ² = 1.69, Potsdam ² = 1.5, Vestfold ² = 0.4, Buskerud ² = 5.62, Kilchberg ² = 0.35, Lausanne ² = 3.32, Canberra ² = 5.29, Massachusetts ² = 3.13), suggesting that the general conclusion of anisoplethy in tristylous L. salicaria is inappropriate.This is Scientific Journal Series Paper Number 19 128 of the Minnesota Agricultural Experiment Station     

Effects of domestication and agronomic selection on phytoalexin antifungal defense in <Emphasis Type="Italic">Phaseolus</Emphasis> beans

Systems of wild and cultivated relatives are good experimental systems to test chemical defense theory because they provide closely related varieties that differ in discrete traits. To determine the relationship between resistance and chemical defense diversity among wild, landrace and cultivar accessions of Phaseolus vulgaris, we measured resistance to fungal infection in laboratory and field experiments, quantified phytoalexin diversity, and assessed the effectiveness of phytoalexin mixtures extracted from live tissue. Results show a gradient of resistance to fungal infections between wild, landrace and cultivar varieties. In the laboratory, wild seedlings were more resistant (93% non-infected) than landrace seedlings (80% non-infected) and modern cultivar seedlings (68% non-infected). Under field conditions wild seedlings were more resistant (97% non-infected) than cultivar seedlings (71% non-infected). Wild seedlings presented the highest phytoalexin diversity (H=1.11), while those of the landrace presented an intermediate level (H=0.97) and cultivar seedlings presented the lowest diversity (H=0.93). No differences were found in total concentrations. The in vitro inhibitory properties on hyphal growth of the isoflavonoid mixtures produced by individual seedlings showed a similar trend. Our results are consistent with similar gradients in other species of Phaseolus beans and resistance to Colletotrichum sublineolum in sorghum.     

Causes,proximate and ultimate

Within evolutionary biology a distinction is frequently made between proximate and ultimate causes. One apparently plausible interpretation of this dichotomy is that proximate causes concern processes occurring during the life of an organism while ultimate causes refer to those processes (particularly natural selection) that shaped its genome. But ultimate causes are not sought through historical investigations of an organisms lineage. Rather, explanations referring to ultimate causes typically emerge from functional analyses. But these functional analyses do not identify causes of any kind, much less ultimate ones. So-called ultimate explanations are not about causes in any sense resembling those of proximate explanations. The attitude, implicit in the term ultimate cause, that these functional analyses are somehow superordinate to those involving proximate causes is unfounded. Ultimate causes are neither ultimate nor causes.     

Shoot tips of wheat as an alternative source for regenerable embryogenic callus cultures

Shoot tips of Triticum aestivum L. cvs. Turbo and Nandu, both summer wheat varieties, were excised from 4 and 10 day-old seedlings, and used for induction of embryogenic callus. A modified L3 medium, supplemented with 10 M 2,4-dichlorophenoxyacetic acid (2,4-d) for culture initiation, and 5 M 2,4-d for subculturing, was optimal; 90% of 4 day-old Turbo seedlings formed embryogenic callus. Optimal plant regeneration was achieved from callus incubated on a modified MS medium without 2,4-d, but supplemented with 2.22 M 6-benzylaminopurine and 0.27 M naphthaleneacetic acid. Plantlets formed via embryogenesis from all embryogenic Turbo calli initiated from 4 day-old explants, with a mean number of 8 regenerants per explant. Regeneration occured via embryogenesis only. Results obtained using Nandu were within the same range.Abbreviations BA 6-benzylaminopurine - 2,4-d 2,4-dichlorophenoxyacetic acid - NAA naphthaleneacetic acid     

Influence of external factors on callose and cellulose synthesis during incubation in vitro of intact cotton fibres with [14C]sucrose

Seed clusters, with adhering fibres, from individual locules of 36-d-old fruit capsules of Gossypium arboreum L. were fed with [¹⁴C]sucrose in vitro. The fibres synthesised, under standard conditions, (13)--D-glucan (callose) and (14)--D-glucan (cellulose) in the ratio of approx. 2:1. Under a great variety of different conditions this product ratio remained more or less constant, even when total glucan synthesis was strongly inhibited with 2,4-dinitrophenol or phloretin, or when stimulated with abscisic acid. In attempts to favour cellulose synthesis, no conditions were found where the ratio was substantially reduced. On the other hand, the ratio could be appreciably increased by inhibiting cellulose synthesis, e.g. with 2,6-dichlorobenzonitrile or coumarin, by anionic detergents such as sodium dodecyl sulphate, by low temperatures, or by increasing the osmotic strength of the incubation medium up to conditions causing plasmolysis. Specific degradation of callose, during incubation of the seed clusters, by exogenous exo-(13)--D-glucanase significantly diminished incorporation of radioactivity into cellulose.Abbreviations DMSO dimethyl sulphoxide - EDTA ethylenediaminetetraacetic acid