An elongated segment of DNA observed between two human α globin genes

Summary Detailed restriction enzyme analysis of the DNA from a Chinese female showed that one of her chromosomes had a >17.5 kb deletion of DNA, including the , 2, and 1 globin genes, which is present in many Southeast Asians with an -thalassemia-1 chromosome. Her normal chromosome had the expected cluster of -like globin genes (5----2-1-3), but the segment of DNA between the two globin genes was elongated by some 0.5–0.7 kb. Analyses of various restriction sites suggested that this normal variant of the human globin gene complex is due to a crossover between a normal chromosome with () and a chromosome with an -thalassemia-2 (–^3.7) and an -21-hybrid gene.     

Transformation of Atriplex gmelini plants from callus lines using Agrobacterium tumefaciens

Atriplex gmelini plants were regenerated via organogensis from hypocotyl explants. Callus lines were induced from the hypocotyl explants on Linsmaier and Skoog (LS) medium supplemented with 1 M benzyladenine and 5 M -naphthaleneacetic acid in the dark. Shoots were regenerated from the callus lines on LS medium supplemented with 20 M thidiazuron and 0.1 M -naphthaleneacetic acid under a high-intensity light condition (450 mol m^–2 s^–1). The regenerated shoots were rooted on LS medium without growth regulators to obtain fully developed plants. We succeeded in transforming Atriplex gmelini from callus lines using Agrobacterium tumefaciens.     

A theoretical investigation into the lipid interactions of m‐calpain

The protease, mcalpain, has been implicated in a number of pathological conditions. The enzyme is a calciumdependent heterodimer whose activity appears to be modulated by membrane interaction involving a segment, TAMRIL, located in domain V of the protein's small subunit. Based on a sequence analysis of mcalpain, using DWIH and hydrophobic moment plot based methodologies, we have shown that this segment may contribute to a lipid interactive, oblique orientated, helical region. Our results could form a basis for future studies on the postulated lipid modulation of mcalpain activity.     

Effects of GABA antagonists,SR 95531 and bicuculline,on GABAA receptor-regulated chloride flux in rat cortical synaptoneurosomes

Synaptoneurosomes isolated from cerebral cortices of male Sprague-Dawley rats were used for studying GABA_A receptor-regulated chloride influx. The in vitro effects of GABA antagonists, SR 95531 (a pyridazinyl GABA derivative) and bicuculline, on pentobarbital-stimulated, muscimol-stimulated or flunitrazepam-enhanced, muscimol-stimulated chloride uptake were studied. The chloride uptake was determined at 30°C, for 5 sec. Pentobarbital and muscimol produced a maximal stimulation of chloride uptake in cortical synaptoneurosomes at 500 M and 50M, respectively. SR 95531 as well as bicuculline had no effect on the basal uptake of chloride. Whereas, SR 95531 (0.3–30 M) and bicuculline (0.1–100 M), when added 5 min before muscimol (50 M), produced a significant concentration-dependent inhibition of muscimol (50 M)-stimulated chloride uptake (IC₅₀ s of 0.89±0.11 M and 13.45±2.10M, respectively). In studies of the inhibitory effects of SR 95531 and bicuculline on pentobarbital (500 M)-stimulated chloride uptake, the IC₅₀ s were 0.81±0.12 M and 3.86±1.14 M, respectively. SR 95531 exhibited a more potent inhibitory effect than bicuculline on flunitrazepam-enhanced, muscimol-stimulated chloride uptake. The results revealed that SR 95531 has a more potent antagonistic effect than bicuculline on GABA_A-regulated chloride flux.     

Organogenesis versus embryogenesis from long-term suspension cultures of cucumber (Cucumis sativus L.)

Suspension cultures were initiated from leaf explant-derived callus of cucumber,Cucumis sativus cv. Hokus, and maintained under two different conditions; (I) continuously in medium with 5 M 2,4-D + 5 M BA, and (II) alternately three cultures in medium containing 5 M NAA + 5 M BA and one culture in 5 M 2,4-D + 5 M BA. After plating on solid medium with 0.5 M KIN + 0.1 M IAA, suspension aggregates from long-term culture in medium with 2,4-D developed into callus, and subsequently formed somatic embryos. These embryos, however, hardly developed into plants. They showed growth arrest and several structural abnormalities. In contrast, organogenesis took place when suspension aggregates from NAA containing medium were plated on solid medium with 0.5 M KIN + 0.1 M IAA. Numerous adventitious buds were regenerated, which quite normally developed into plants. Sucrose at low concentration of 1% improved plant formation. On the average thirty complete plants were obtained from each ml of suspension. It is discussed why adventitious buds develop into plants so well, whereas somatic embryos are prone to growth arrest and abnormal development.Abbreviations BA 6-benzylaminopurine - KIN kinetin - IAA indole-3-acetic acid - NAA 1-naphthaleneacetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid     

Ganglioside-cholera toxin interactions: a binding and lipid monolayer study

Summary On t.l.c. plates ¹²⁵I-cholera toxin binds to a disialoganglioside tentatively identified as GD_lb with about 10 times less capacity than to ganglioside GM₁. Binding of labeled toxin to both gangliosides was abolished in presence of excess amounts of unlabeled B subunit. Ganglioside extracts from human or pig intestinal mucosa showed toxin binding to gangliosides GM₁ and GD_1b. In ganglioside-containing lipid monolayers the penetration of the toxin was independent of the ganglioside binding capacity.Abbreviations GM₂ Gal-NAc14Gal(3-2NeuAc)14G1c1Cer - GM₁ Gal3Ga1-NAc14Gal(32NeuAc)14G1c11Cer - GD_1a NeuAc23Ga113Gal-NAc14Gal(32NeuAc)14G1c11Cer - GD1b Gall3Gal-NAcl4Gal(32NeuAc82NeuAc)14Glc11Cer - GT_1b NeuAc23Ga113Ga1-NAcal4Gal(3-2NeuAc82NeuAc)14G1c11Cer - dpPC 1,2-hexadecanoyl-sn-glycero-3-phosphocholine - dpPE 1,2-hexadecanoyl-sn-glycero-3-phosphoethanolamine     

Problems in estimating growth rates of marine phytoplankton from short-term14C assays

Growth rate estimates () of phytoplankton populations that were sampled from nitrogen-limited continuous cultures and then incubated for short durations in batch culture with added¹⁴C-HCO₃ ^– were significantly different than steady-state growth rates () for 3 of 5 marine phytoplankton species. Two diatoms,Thalassiosira weissflogii andChaetoceros simplex, displayed virtually identical growth rates (=) over a wide range of, whereas for a third diatom,Phaeodactylum tricornutum, was overestimated by an average of 40% compared to. In contrast, was underestimated by the¹⁴C technique for the two remaining species: up to 40% at a steady-state of 1.0 day^–1 for the chlorophyteDunaliella tertiolecta and up to 100% at of 1.4 day^–1 for the haptophytePavlova lutheri. For the latter two species the divergence between and appeared to increase with increasing steady-state. A simple model of labeled and total carbon flow between the aqueous phase and cellular biomass was constructed to demonstrate that respiration was negligible when=, but was significant when>. In the cases in which<, a rapid physiological alteration presumably took place once the steady state was disturbed and cells were placed in the incubation chambers, which perhaps was related to the nutritional state of the cultures at the time of sampling. Questions thus are raised regarding our ability to measure accurately primary productivity from shipboard experiments with confined samples of phytoplankton from nutrient-impoverished waters that probably are less hardy than the laboratory cultures used in these studies.     

Polarities in Mammalian Evolution Seen Through Homologs of the Inner Cell Mass

Embryogenetic pathways differ markedly among monotremes, marsupials, and placentals, and their analysis provides information of fundamental importance to recognition of mammalian evolutionary directions. The cap of cuboidal cells of the marsupial late unilaminar blastocyst, generally known as the embryonic area, probably is induced to form (prior to origin of Hensen's node) by signals from earliest hypoblastic cells (anterior visceral endoderm). The thickened cap is a medullary plate of sauropsid terminology because it includes epiblastic cells presumptive to neurectoderm (including neural crest), Hensen's node, primitive streak, and gut endoderm. The remainder of the definitive embryo (i.e., parts of epidermal origin, including ectodermal placodes) derives from squamous ectoderm (surrounding the medullary plate) of the blastocyst's ill-named trophoblastic area. Amniotic ectoderm develops farther distally within the trophoblastic area. The autapomorphic inner cell mass (ICM) of placental mammals is homologous to medullary plate of the marsupial blastocyst plus morphologically undefined, proximal parts of surrounding ectoderm (of the trophoblastic area). Considerations of early cell lineages in marsupials are greatly affected by recognition that the boundary between future embryonic and extra-embryonic tissues does not match the margin of the medullary plate (i.e., embryonic area). Marsupials and monotremes largely conform to sauropsid early embryogenesis, but placentals express, at earliest developmental stages, innovations unique within Amniota that are linked to early establishment of the brain. Neonatal marsupials and hatchling monotremes are extremely altricial and closely comparable anatomically/physiologically; they share a temporal pattern in combining early morphogenesis of craniofacial features (related to suckling) with deferral of telencephalic completion into postnatal/posthatching life. Placentals contrast greatly in establishing the central nervous system prior to rudiments of the cranial skeleton and associated musculature, and they complete essentials of forebrain development before birth. Comparative evidence from transitory periderm suggests that primordial eutherians had extremely altricial hatchlings or newborns, whichever was the mode of early development. Details remain unknown about the origin of the unique specialization of ICM plus encapsulating trophoblast from the more generalized blastula of ancestral synapsids.     

Carotenoid pigments of Stigmatella aurantiaca (Myxobacterales)

Summary In this first article on the carotenoids of Myxobacterales we report on the minor carotenoids of Stigmatella aurantiaca: phytoene, phytofluene, lycopene, -carotene, 4-keto--carotene, 1,2-dihydro-1-hydroxy--carotene, 4-keto-1,2-dihydro-1-hydroxy--carotene, 4-keto-1,2-dihydro-1-hydroxy-torulene, and 1,2,1,2-tetrahydro-1,1-dihydroxy-lycopene. These pigments account for about 10% of total carotenoids.     

Activation of LA-N-2 Cell Phospholipase D by Amyloid Beta Protein (25–35)

Amyloid protein is the major protein component of neuritic plaques found in the brain of Alzheimer's disease. The activation of phospholipase D by amyloid beta protein (25–35), quisqualate and phorbol 12, 13-dibutyrate was investigated in LA-N-2 cells by measuring phosphatidylethanol formation. The activation of phospholipase D by quisqualate and AP (25–35) was calcium-independent. The AP (25–35) and quisqualate activation of phospholipase D appeared to be mediated through a pertussis toxin-sensitive GTP-binding protein. Phospholipase D activation by AP (25–35), quisqualate and phorbol dibutyrate was not blunted by the protein kinase C inhibitors, staurosporine, H-7 and RO-31-8220. However, it was abolished by overnight exposure to phorbol dibutyrate. This activation of phospholipase D was prevented by the tyrosine kinase inhibitor, genistein but not by tyrophostin A. Several excitatory amino acid antagonists were tested for their ability to prevent the phospholipase D activation by quisqualate and AP (25–35). Only NBQX was effective with an IC₅₀ of 75 M for AP (25–35) and quisqualate. Activation of phospholipase D by AP or quisqualate was absent in LA-N-2 cells previously desensitized by quisqualate or AP (25–35), but the activation by phorbol dibutyrate was unaltered. The responsiveness to AP and quisqualate in previously desensitized cells reappeared subsequent to a period of resensitization. The observations with the antagonist NBQX, and the desensitization and resensitization experiments, are consistent with a receptor occupancy mediated activation of phospholipase D by quisqualate and by AP (25–35).     

Heterochromatin variation in Cryptobothrus chrysophorus

Northern and southern races of the Australian grasshopper Cryptobothrus chrysophorus share the same diploid chromosome number (2n=23, 24). The northern race is differentiated from the southern by fixed extra blocks of heterochromatin located distally on five of the six medium pairs of autosomes (M4, 5, 6, 8 and 9). The megameric M7 pair, which is completely heterochromatic in both races, is also frequently larger in the northern race. Additionally, while there is considerable polymorphism for the presence of supernumerary heterochromatic segments on the two smallest autosome pairs (S10, 11) in both races, the precise character of this polymorphism is strikingly different between them. That found in the north is both more extensive and more variable. An analysis of the patterns of C-banding obtained in neuroblast c-mitoses indicates even more variation within and between races than was anticipated from the patterns of heteropycnosis seen at first prophase of male meiosis. Thus, while the distal blocks on the M4, 6, 8 and 9 elements in the northern race invariably C-band those of the M5 never do. On the other hand polymorphisms for C-bands on the M5, 6, 8 and 9 are seen in some populations of the southern race but in regions which are not visibly heteropycnotic at meiosis. Polymorphisms for the pattern of C-banding also occur in the northern race populations in the M4, 6, 8 and 9 elements and some of these are associated with clear length differences in the chromosomes concerned. Others involve differences in the expression of the distal C-bands in M8 and 9 which vary from dark to intermediate. The supernumerary segments on the S10 and 11 pairs are especially variable in respect of their C-banding properties. Some are entirely C-banded, some show no C-banding whatsoever and some are composed of both banded and unhanded regions. Banding is again most pronounced, however, in the northern race. Finally the character of the megameric M7 is strikingly different in the two races not only in respect of its size, which is sometimes larger than that of the south, but also in respect of the extent of C-banding which is always more complex in the northern form irrespective of its size.     

The NGF complex from the African ratMastomys natalensis

The 7S NGF complex from the male mouse submaxillary gland consists of the , and subunits in the ratio ₂₂. The (NGF) subunit contains all the known biolocial activity of 7S NGF. The and subunits are both members of glandular kallikrein gene family, yet only subunit has protease activity. The subunit plays a role in the processing of preproNGF to its mature form, while the role of the subunit is not yet understood. Despite the fact that 7S NGF has been extensively characterized, no other NGF complex has been characterized, nor have the or subunits been observed in tissues which express NGF. We have therefore purified and characterized the NGF complex from the submaxillary glands of the ratMastomys natalensis in order to more fully understand the roles of the and subunits. The NGF complex from M. natalensis contains subunits similar to those found in mouse 7S NGF. Although similar, there are significant differences between mouse and M. natalensis NGF complexes, especially in the degree of post-translational modification of the and NGF subunits, the expression of esterase activity and the ease with which the complexes dissociate. Evidence is presented that suggests that the NGF complex from M. natalensis may consist of subunits in the ratio ₂. The amino acid sequence of the M. natalensis NGF suggests some, but not all, ways in which these differences arise.Special issue dedicated to Dr. Lawrence Austin     

Response to Striefel and Glaros

In this response to the comments of Drs. Striefel and Glaros, we take issue with Dr. Striefel's assertion that the Declaration of Helsinki should not be regarded as a core ethical document, and demonstrate that his claim that the Declaration has no significance in or recognition by agencies of the United States Federal Government is in error. A reading of FDA and DHHS documents shows that the Declaration is not, as Dr. Striefel suggested, only an aspirational document, but also is clearly regarded as mandatory. Dr. Glaros' observations regarding the slippery concept of efficacy is very much on point, and we certainly agree with his call for more data. It is the means by which that can be accomplished that is problematic. We have suggested that the logic of placebo controls developed to render drug efficacy studies more rigorous and scientifically grounded does not translate well to many psychological and behavioral studies of efficacy. It is important for our discipline to rethink the standards by which new behavioral and psychological (including psychophysiological) interventions can be demonstrated to be efficacious. One possible approach that avoids ethical pitfalls and design impossibilities is the active control, which requires good demonstration of assay sensitivity. We hope that this discussion may stimulate further discussion about other approaches that do not depend upon the flawed placebo orthodoxy.     

Evidence of β-carotene 7,8(7′,8′) oxygenase (β-cyclocitral,crocetindial generating) in Microcystis

A -carotene oxygenase is described which occurs in the Cyanobacterium Microcystis. It cleaves -carotene and zeaxanthin specifically at the positions 7,8 and 7,8, while echinenone and myxoxanthophyll are not affected. The oxidative cleavage of -carotene leads to the formation of -cyclocitral and crocetindial and that of zeaxanthin to hydroxy--cyclocitral and crocetindial in nearly stoichiometric amounts. Oxidant is dioxygen as has been demonstrated by high incroporation (86%) of ¹⁸O₂ into -cyclocitral. -Carotene oxygenase is membrane bound, sensitive to sulfhydryl reagents, antioxidants and chelating agents. Iron seems to be an essential part of the enzyme activity. Cofactors necessary for the reaction could not be detected.Abbreviations TLC thin layer-chromatography - PIPES piperazine-N,N-bis-(2-ethanesulfonate) Na - TES 2{[tris-(hydroxymethyl)-methyl]-amino} ethanesulfonic acid Dedicated to Professor G. Drews on occasion of his 60th birthday     

Pacemaker synaptic interactions: Modelled locking and paradoxical features

This communication describes a model for two pacemaker (i.e., regularly firing) nerve cells, such that one elicits IPSP's in the other. The assumptions involve essentially a linear dependence (delay function) of the postsynaptic interval lengthening (or delay) produced by the IPSP's on the position (or phase) with respect to the preceding spike of the latter's arrival. When the number of IPSP's in an interval increases, both the slope and intercept of the delay function increase, the former remaining under 2 and the latter unboundedly. Assumptions are more or less close to the actual biological reality, or are made for convenience. A recurrence equation for the phase can be calculated, as well as an expression for the locking phase (see below). Plots of postsynaptic vs presynaptic firing intensity averaged over steady conditions, e.g. of mean rates or intervals, are formed by a sequence of relatively broadparadoxical segments exhibiting positive slopes 1, 2, 1/2, 3, 1/3, ..., indicating that inhibited discharges are made more intense by those increases in inhibitory arrivals. These segments are separated by narrower intercalated segments where behavior is unclear except for a large overall negative slope, indicating that inhibited discharges are weakened markedly by other increases in inhibitory arrivals. Across the successive paradoxical segments that correspond to more and more intense presynaptic discharges (i.e., to higher rates or shorter intervals), postsynaptic intensities, though overlapping in part, become weaker and weaker. At the extremes, when the presynaptic discharge is very weak, or very intense, the postsynaptic cell tends to its natural undisturbed firing, or to not firing at all, respectively. The pre- and postsynaptic discharges inevitably achieve eventually an invariant relation, i.e., will lock at a constant phase, regardless of the phase of the first IPSP arrival. The characteristics of this behavior (e.g., the rate bounds of the paradoxical segments, or the magnitude of the locked phase) depend on such givens as presynaptic and postsynaptic pacemaker rates or intervals, and as the slope or intercept of the delay function.     

Movements and diurnal behaviour patterns in a Baltic grey seal (Halichoerus grypus)

A sub-adult grey seal was caught and tagged with a satellite relay data logger at Tihällan (61°2800N, 17°2540E), Sweden, on 21 August 1992. We tracked it for 115 days during which time we recorded 841 locations, data on 3556 dives and 80 haulout periods. It travelled 730 km in the first 12 days to the southern Archipelago Sea. It stayed most of the remaining time within 25 km of a haulout site, with a major exception of an 8-day trip of 520 km. Dive shape and swimming activity were used to distinguish dive types. Square-shaped foraging dives were more common over the midday period. Haulout periods were more common during the night.     

Biochemische Untersuchungen zur Frage der Existenz eines “silent Gene” im Polymorphismus der Pseudocholinesterasen

Zusammenfassung Zwei Seren des Phänotypus Ch₁SS, die unter Standardtestbedingungen keine Pseudocholinesteraseaktivität aufwiesen, wurden mit Hilfe der Stärkegel-Elektrophorese, der Mikromanometrie und mit immunologischen Methoden näher untersucht.Nach elektrophoretischer Auftrennung läßt sich in diesen silent gene-Seren eine Pseudocholinesteraseaktivität nachweisen; im Vergleich zu der Aktivität von Normalserum erscheint diese sehr gering und läßt sich nur in der c₄-Zone erkennen. Die Identifizierung gelingt mit Hilfe spezifischer Färbeverfahren unter Verwendung verschiedener Substrate und Inhibitoren.Die quantitative Bestimmung von Pseudocholinesteraseaktivität im silent gene-Serum mit mikromanometrischen Methoden ergab eine Aktivität von 2–3% gegenüber den Kontrollen (Benzoylcholin als Substrat).Durch Immunisierung von Kaninchen mit gereinigtem Pseudocholinesteraseprotein wurden Antiseren erhalten; zwischen silent gene-Serum und diesen Antiseren konnten Präzipitationsreaktionen im Immuno-Diffusionstest, in der Immuno-Elektrophorese und mit einer Immuno-Adsorptionsmethode nachgewiesen werden. Diese Ergebnisse lassen annehmen, daß bei den von uns untersuchten Fällen das silent gene im Pseudocholinesterasepolymorphismus eine Enzymproteinsynthese steuert; das nachgewiesene Pseudocholinesteraseprotein scheint sich qualitativ von dem Enzymprotein des Normalserums zu unterscheiden.

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Two sera without pseudocholinesterase activity corresponding to the homozygous phenotype Ch₁SS are examined by electrophoretical, manometric, and immunological methods.These silent gene sera show no activity under the common conditions (spectrophotometric assay).After electrophoretical separation of silent gene serum an esterase activity is found which can be identified as pseudocholinesterase activity, although it is weak in comparison with the activity of usual sera. The pseudocholinesterase activity of silent gene serum can be demonstrated only in the zone c₄ where 90% of the total activity is present if usual serum is inserted. The identification has been achieved by staining procedures applying several substrates and inhibitors.Quantitative estimation of this pseudocholinesterase activity was carried out by micromanometric assays with benzoylcholine as substrate. The activity of silent gene sera was 2–3% of normal serum.Antisera against human pseudocholinesterase-protein have been obtained by immunization of rabbits with a highly purified enzyme protein. Between these antisera and the homozygous silent gene sera precipitates were found in immuno double-diffusion tests and immunoelectrophoresis. They could be identified as pseudocholinesterase protein by esterase staining under various conditions.Quantitative estimations have been carried out by immuno-adsorption assays comparing the amount of antibody fixed by usual serum and by silent gene serum.The results presented in this paper suggest that the silent gene in pseudocholinesterase polymorphism induces in these two cases the synthesis of an enzyme protein which is similar to the enzyme protein of usual pseudocholinesterase. The weak activity is due to a qualitative difference between silent gene enzyme protein and the normal pseudocholinesterase protein. A structural alteration of the enzyme protein is assumed to be more likely than a quantitative difference in protein synthesis.

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Einige der Ergebnisse dieser Arbeit wurden auf dem 2nd Meeting of the Federation of European Biochemical Societies, Wien 1965, vorgetragen.

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Wesentliche Teile dieser Arbeit werden von R. A. Hofmann der Medizinischen Fakultät der Universität Freiburg i. Br. als Inauguraldissertation vorgelegt.     

Insertion of IS1 into the rpsE gene for ribosomal protein S5 causes cold-sensitivity in Escherichia coli

Summary The nucleotide sequence of the rpsE gene for the ribosomal protein S5 of a mutant of E. coli showing cold-sensitive growth revealed the presence of an insertion sequence, IS1, near the 3-end of the gene. This mutant grows very slowly even at the permissive temperature (30°C). At temperatures below 20°C, its growth becomes negligible. It is likely that the presence of IS1 disturbs the continued translation of the polycistronic messenger RNA of the spc-operon from the point of IS1 insertion downwards, especially at the unfavorable temperatures.     

Axillary shoot induction and plant regeneration in Plantago ovata Forssk

Axillary shoot induction and plant regeneration were obtained in Plantago ovata. The optimum medium for inducing axillary shoots was Murashige & Skoog (MS) medium [5] supplemented with 4.6 M kinetin and 0.05 M NAA. Rooting of shoots was best on half-strength MS medium containing 5.0 M IBA and 0.05 M kinetin. The regenerated plants were similar to the control plants in karyotypic and phenotypic details.     

Concluding remarks

On the basis of symposium contributions onChlorella, Hibbertia, Eucalyptus, Ambrosia and on numerical approaches some fundamental problems of (bio)systematics, evolution, and taxonomic categories are discussed: Methods available for analysing affinities; conflicting evidence from phenetic, biochemical, cytogenetic and other analyses; further classification problems in cases of intermediacy, etc. While sibs of various levels and their natural hierarchy often can be objectively defined, this appears impossible for particular taxonomic levels itself (e. g. species). A single objective taxonomic system of organisms is unrealistic. Certain guiding lines for relative and practicable concepts of species and genus are proposed.Presented at the symposium Speciation and the Species Concept during the XIIth International Botanical Congress, Leningrad, July 8, 1975.