The protegulum and juvenile shell of a Recent articulate brachiopod: patterns of growth and chemical composition

Patterns of shell formation and the chemical composition of the shell deposited during early post-larval life were investigated in laboratory-reared cultures of the Recent articulate brachiopod Terebraralia transversa (Sowerby). A non-hinged protegulum averaging 148 pm in length is secreted by the mantle within a day after larval metamorphosis. The inner surface of the protegulum exhibits finely granular, non-fibrous material. A rudimentary periostracum constitutes the outer layer of the primordial shell. and concentrically arranged growth lines are lacking. By four days post-metamorphosis, a brephic type of juvenile shell develops from periodic additions of shell material to the anterior and lateral edges of the protegulum. Imbricated secondary fibers occur throughout the inner layer of the newly formed juvenile shell, and a rudimentary hinge apparatus is present posteriorly. The external surface of the shell exhibits concentric growth lines anterior to the caudally situated protegulum, and unbranched punctae begin to form in the subperiostracal region of the shell. At 23 days post-metamorphosis, the shell weighs an average of 1.7 μg and measures 318 μm in length. Electron microprobe analyses reveal that the protegulum is calcified. Minor amounts of sulfur, magnesium, iron, chlorine, aluminum, and silicon are also present in protegula and juvenile shells. Based on electron diffraction data, the mineral phase of juvenile shells consists of calcite, and protegula also appear to contain calcite.     

The ontogeny of shell secretion in Terebratalia transversa (Brachiopoda, Articulata). II. Formation of the protegulum and juvenile shell

The fine structure of the shell and underlying mantle in young juveniles of the articulate brachiopod Terebratalia transversa has been examined by electron microscopy. The first shell produced by the mantle consists of a nonhinged protegulum that lacks concentric growth lines. The protegulum is secreted within a day after larval metamorphosis and typically measures 140-150 micron long. A thin organic periostracum constitutes the outer layer of the protegulum, and finely granular shell material occurs beneath the periostracum. Protegula resist digestion in sodium hypochlorite and are refractory to sectioning, suggesting that the subperiostracal portion of the primordial shell is mineralized. The juvenile shell at 4 days postmetamorphosis possesses incomplete sockets and rudimentary teeth that consist of nonfibrous material. The secondary layer occuring in the inner part of the juvenile shell contains imbricated fibers, whereas the outer portion of the shell comprises a bipartite periostracum and an underlying primary layer of nonfibrous shell. Deposition of the periostracum takes place within a slot that is situated between the so-called lobate and vesicular cells of the outer mantle lobe. Vesicular cells deposit the basal layer of the periostracum, while lobate cells contribute materials to the overlying periostracal superstructure. Cells with numerous tonofibrils and hemidesmosomes differentiate in the outer mantle epithelium at sites of muscle attachments, and unbranched punctae that surround mantle caeca develop throughout the subperiostracal portion of the shell. Three weeks after metamorphosis, the juvenile shell averages about 320 micron in length and is similar in ultrastructure to the shells secreted by adult articulates.     

Ontogenetic phase shifts in metabolism: links to development and anti-predator adaptation

The allometric relationships between resting metabolism (VO₂) and body mass (M), VO₂ = a_iM^b, are considered a fundamental law of nature. A distinction though needs to be made between the ontogeny (within a species) and phylogeny (among species) of metabolism. However, the nature and significance of the intraspecific allometry (ontogeny of metabolism) have not been established in fishes. In this study, we present experimental evidence that a puffer fish ranging 0.0008–3 g in wet body mass has four distinct allometric phases in which three stepwise increases in scaling constants (a_i, i = 1–4), i.e. ontogenetic phase shifts in metabolism, occur with growth during its early life stages at around 0.002, 0.01 and 0.1 g, keeping each scaling exponent constant in each phase (b = 0.795). Three stepwise increases in a_i accompanied behavioural and morphological changes and three peaks of severe cannibalism, in which the majority of predation occurred on smaller fish that had a lower value of a_i. Though fishes are generally highly fecund, producing a large number of small eggs, their survivability is very low. These results suggest that individuals with the ability to rapidly grow and step up ‘a_i’ develop more anti-predator adaptation as a result of the decreased predatory risk.     

Sediment particles as a cause of nacre staining in the freshwater mussel,Amblema plicata (Say) (Bivalvia: Unionidae)

Brown stains sometimes appear in the inner shell layers (nacre) of freshwater mussels. An electron microprobe was used to analyze the stained nacre of the unionid Amblema plicata (Say, 1817) from selected localities on the Mississippi River in the vicinity of LaCrosse and Prairie du Chien, Wisconsin.Several elements such as Na, Mg, Al, Si, P, S, Cl, K, and Fe are more highly concentrated in stained than in unstained nacre. Concentrations of these elements relative to Ca were found to vary significantly among the localities from which the specimens were obtained. Ratios have significantly higher variances downstream of the confluence of the Yellow and Mississippi Rivers, downstream of a barge fleeting area, near the town of Marquette, Iowa, near the site of sewage effluent for Prairie du Chien, Wisconsin and downstream of a scrap metal yard near LaCrosse, Wisconsin in contrast to control localities.Suspended silt, the result of runoff and river activity (barge traffic, dredging, pleasure boating) may be the stimulus for stain formation. Clay minerals adsorb accessory elements and in turbid water are trapped within the pallial space of A. plicata. The mussel secretes an organic-rich, periostracum-like layer over the entrapped sediment, and later reverts to prismatic and finally nacreous shell deposition. Some of the elements found in the stain could directly disturb Ca metabolism by competing with Ca for binding sites in shell aragonite.     

Age and distribution of the Late Devonian brachiopod genus Dzieduszyckia Siemiradzki, 1909 in southern China

Conodont biostratigraphical work was done at four sections recently found with occurrence of the rhynchonellide brachiopod genus, Dzieduszyckia Siemiradzki, in southern Guangxi and in the border area between Dushan County of Guizhou and Nandan County of Guangxi, South China. These sections represent two different types of facies, i.e., carbonate platform and intraplatform basin. The conodont analysis reveals that this genus occurs in the Upper triangularis Zone and the Middle crepida Zone at the Dazhai Section, through the Lower to Upper crepida zones at Dalong, and is restricted within the Upper rhomboidea Zone at the two intraplatform basin sections (Changtang and Duli). This result demonstrates that the occurrence of these peculiar rhynchonellide brachiopods in South China, regardless of the depositional environments, is within the Lower Famennian instead of the previously suggested Upper Famennian. Furthermore, this brachiopod genus in South China began to inhabit on the carbonate platform almost since the beginning of the Famennian and did not extend to the intraplatform basin facies until the late Early Famennian.Available biostratigraphic data indicate that during the Early and Middle Famennian, Dzieduszyckia is widely distributed not only in South China, but also throughout the world, such as Morocco and southern Ural.Observation on the new collections from the four studied sections reveals that the peculiar rhynchonellide brachiopods have a great morphological variation within each section. Significant differences existed among the collections from different sedimentary settings and localities, probably reflecting the environmental and geographic constraint on the morphology of Dzieduszyckia. Samples from different layers in the same section have nearly identical morphological variation, suggesting the temporal inheritance in morphology of the rhynchonellide brachiopod.     

Copper and zinc metabolism with solid tumor growth

The variation in copper and zinc metabolism with tumor growth appears to relate directly to progression or regression of the disease. Historically, elevations in serum copper have been used as clinical indicators in hematological neoplasms since the early 1960s. More recently, we have monitored breast, colo-rectal, and lung cancer patients for a six-month period through courses of cytotoxic chemotherapy to determine copper and zinc changes with tumor growth. Groups were divided into responders and nonresponders blind to their serum copper and zinc levels. Trends in elevated serum copper with active disease have shown similar trends in decreasing values with effective therapy, but normalization was at a slower rate. Serum zinc levels in the same patients were markedly below normal and did not increase in the study period. The clinical significance or elevated serum copper and depressed serum zinc is discussed and the potential relationship between the two elements is explored. A solid tumor-bearing rat model, mammary adenocarcinoma R 3230 AC, has detailed more of the changes in copper and zinc metabolism with solid tumor growth. Serum copper and zinc varied with tumor mass, as in clinical studies. Liver values of the two essential metals did not change significantly, but liver-related copper-containing enzymes showed marked variations. Ceruloplasmin in serum increased with increasing tumor mass, as would be expected with the increased serum copper levels. Cytochrome c oxidase activity in liver homogenates from tumor-bearing animals was significantly depressed.     

A protein involved in co-ordinated regulation of inorganic carbon and glucose metabolism in the facultative photoautotrophic cyanobacteriumSynechocystis PCC6803

The involvement of a gene ofSynechocystis PCC6803,icfG, in the co-ordinated regulation of inorganic carbon and glucose metabolism, was established. TheicfG gene codes for a 72 kDa protein, which shows no homology with those registered in data libraries. Expression oficfG required glucose, the actual inducer probably being glucose-6-phosphate, and was independent of light and of the external inorganic carbon concentration. Mutants carrying an inactivated copy oficfG were constructed. Their growth characteristics were identical to those of the wild type under all regimes except in limiting inorganic carbon with glucose being present either before or after the transfer to the limiting conditions. These conditions completely prevented growth, both in the light and in the dark. The inhibition could be relieved by several intermediates of the tricarboxylic acid cycle. Assays of various enzymic activities related to inorganic carbon uptake and to its assimilationvia either the Calvin cycle or phosphoenolpyruvate carboxylase did not reveal the level of action of IcfG. Possible models include a blockage of the assimilation of both carbon sources in the absence of IcfG, or the inhibition of Ci incorporation route(s) essential under limiting inorganic carbon conditions, even when glucose is present, and even in the dark.     

Tissue-specific palatability and chemical defenses against macropredators and pathogens in the common articulate brachiopod Liothyrella uva from the Antarctic Peninsula

The punctate terebratulid brachiopod Liothyrella uva is the most common brachiopod species in Antarctica. Whole brachiopods, either live or freeze dried and ground into a powder and suspended in alginate, were unpalatable to the sympatric macropredators Odontaster validus (an abundant, omnivorous sea star) and Notothenia coriiceps (an abundant, omnivorous, epibenthic fish). The unpalatability of these ground tissues coupled with that of lipophilic extracts of whole L. uva presented in alginate pellets to O. validus, suggests an involvement of chemical defenses. Several isolated brachiopod tissues were also unpalatable to O. validus after being freeze dried, ground and suspended in alginate, but only the pedicle was unpalatable in such preparations to both O. validus and N. coriiceps. This observation is consistent with the Optimal Defense Theory since the pedicle is the only tissue not protected inside the brachiopod shell. There was, however, no correlation between the energetic content and unpalatability of any of the individual tissues. Organic extracts of tissues involved in feeding (lophophore and intestine–stomach) had relatively strong antimicrobial activity when assayed against several strains of Antarctic bacteria. However, the lophophore was palatable to both macropredators, suggesting nonoverlapping chemical defenses are involved in protection against predators and pathogens.     

Succinate metabolism related to lipid synthesis in the housefly Musca domestica L

The metabolism of succinate was examined in the housefly Musca domestica L. The labeled carbons from [2,3-¹⁴C]succinate were readily incorporated into cuticular hydrocarbon and internal lipid, whereas radioactivity from [1,4-¹⁴C]succinate was not incorporated into either fraction. Examination of the incorporation of [2,3-¹⁴C]succinate, [1-¹⁴C]acetate, and [U-¹⁴C]proline into hydrocarbon by radio-gas-liquid chromatography showed that each substrate gave a similar labeling pattern, which suggested that succinate and proline were converted to acetyl-CoA prior to incorporation into hydrocarbons. Carbon-13 nuclear magnetic resonance showed that the labeled carbons from [2,3-¹³C]succinate enriched carbons 1, 2, and 3 of hydrocarbons with carbon-carbon coupling showing that carbons 2 and 3 of succinate were incorporated as an intact unit. Radio-high-performance liquid chromatographic analysis of [2,3-¹⁴C]succinate metabolism by mitochondrial preparations showed that in addition to labeling fumarate, malate, and citrate, considerable radioactivity was also present in the acetate fraction. The data show that succinate was not converted to methylmalonate and did not label hydrocarbon via a methylmalonyl derivative. Malic enzyme was assayed in sonicated mitochondria prepared from the abdomens and thoraces of 1- and 4-day-old insects; higher activity was obtained with NAD⁺ in mitochondria prepared from thoraces, whereas NADP⁺ gave higher activity with abdomen preparations. These data document the metabolism of succinate to acetyl-CoA and not to a methylmalonyl unit prior to incorporation into lipid in the housefly and establish the role of the malic enzyme in this process.     

Characterization of rat lymphocyte primary culture for the development of an in-vitro mutagenesis assay: Effect of interleukin-2 and 2-mercaptoethanol on the activities of intermediary metabolism enzymes and cell proliferation

Efficient energy utilization is essential for cell growth; in an attempt to improve the growth conditions of the rat T-lymphocyte culture model for potential use in studying the mutagenic activity of carcinogens in vitro, we have investigated the effects of phytohemagglutinin (PHA), interleukin-2 (IL-2) and 2-mercaptoethanol (2-ME) on the activities of intermediary metabolism enzymes and cell proliferation. Isolated lymphocytes were cultured in the presence and absence of PHA, IL-2, or 2-ME. The intermediary metabolism enzymes investigated were glutamate dehydrogenase, glutamate-pyruvate transaminase, malate dehydrogenase, isocitrate dehydrogenase, lactate dehydrogenase, pyruvate kinase, and fatty acid synthetase (FAS). Measurable activity of all enzymes investigated, except for FAS, was detected in PHA-stimulated cells cultured with IL-2 or 2-ME. The unstimulated lymphocytes had significantly lower enzyme activity than stimulated cells. The combination of all three agents showed increased enzyme activity. This increase in activity brought about by the combination of the three agents was not reproduced by either agent acting alone. In general, the increase in enzyme activity correlated with cell proliferation as measured by [³H]thymidine uptake in PHA-stimulated cultures containing IL-2 and/or 2-ME. The results suggest that the addition of exogenous IL-2 and 2-ME enhances metabolic function and may be beneficial in in vitro culture of rat lymphocytes.Abbreviations PHA phytohemagglutinin - IL-2 interleukin-2 - 2-ME 2-mercaptoethanol - GDH glutamate dehydrogenase - GPT glutamate-pyruvate transaminase - MDH malate dehydrogenase - ICD isocitrate dehydrogenase - LDM lactate dehydrogenase - PK pyruvate kinase - FAS fatty acid synthetase     

The Arabidopsis sweetie mutant is affected in carbohydrate metabolism and defective in the control of growth, development and senescence

Sugars modulate many vital metabolic and developmental processes in plants, from seed germination to flowering, senescence and protection against diverse abiotic and biotic stresses. However, the exact mechanisms involved in morphogenesis, developmental signalling and stress tolerance remain largely unknown. Here we report the characterization of a novel Arabidopsis thaliana mutant, sweetie , with drastically altered morphogenesis, and a strongly modified carbohydrate metabolism leading to elevated levels of trehalose, trehalose-6-phosphate and starch. We additionally show that the disruption of SWEETIE causes significant growth and developmental alterations, such as severe dwarfism, lancet-shaped leaves, early senescence and flower sterility. Genes implicated in sugar metabolism, senescence, ethylene biosynthesis and abiotic stress were found to be upregulated in sweetie . Our physiological, biochemical, genetic and molecular data indicate that the mutation in sweetie was nuclear, single and recessive. The effects of metabolizable sugars and osmolytes on sweetie morphogenesis were distinct; in light, sweetie was hypersensitive to sucrose and glucose during vegetative growth and a partial phenotypic reversion took place in the presence of high sorbitol concentrations. However, SWEETIE encodes a protein that is unrelated to any known enzyme involved in sugar metabolism. We suggest that SWEETIE plays an important regulatory function that influences multiple metabolic, hormonal and stress-related pathways, leading to altered gene expression and pronounced changes in the accumulation of sugar, starch and ethylene.     

The tracheal supply and muscle metabolism during muscle growth in the puparium of Calliphora vomitoria

Thirty hours after puparium formation in Calliphora, the larval tracheal system is replaced by an air-filled pupal system. This is characterized initially by many tufts of tracheae and coiled tracheoles lying in the blood. Between the third and fourth day, the sixth dorsal longitudinal flight muscles are practically without attached tracheae and their longitudinal growth can partially occur when oxygen uptake is inhibited with potassium cyanide. Sodium iodoacetate prevents muscle growth. After the fifth day of development the pupal tracheoles spread out over the surface of the developing adult muscles. Between the seventh and ninth day, longitudinal growth and increases in the diameter of the myofibrils are halted by cyanide and iodoacetate. Some longitudinal growth and an increase in the total protein content of the muscles can occur in 1% oxygen. Air filling of the adult tracheae takes place 2–3 hr before the emergence of the adult and is accompanied by an increase in oxygen consumption of the thorax. The metabolism and growth of the muscles is discussed with respect to their changing oxygen supply.     

Development of primary culture of ovine fetal hepatocytes for studies of amino acid metabolism and insulinlike growth factors

Summary We report the development and characterization of a system of primary culture of ovine fetal hepatocytes to aid in the understanding of the cellular regulation of fetal growth and metabolism with emphasis on amino acid metabolism and insulinlike growth factor gene expression and to allow comparison to in vivo studies. Hepatocytes were isolated from late gestation fetal lambs by in situ perfusion and collagenase digestion utilizing occlusion of the ductus venosus to limit intrahepatic shunting. Hepatocytes were cultured in media modified to mimic fetal concentrations of glucose, lactate, and amino acids. Ovine fetal hepatocytes in primary culture maintain the pattern of fetal amino acid production and utilization seen across the fetal liver in vivo. Specifically, there is a net production of serine and a net utilization of glycine. Cultured ovine fetal hepatocytes specifically increase tritiated thymidine incorporation in response to insulin and insulinlike growth factor II (IGF-II). IGF-II mRNA abundance is high and IGF-I mRNA is low in cultured ovine fetal hepatocytes as in the fetal sheep liver in vivo. These data demonstrate the successful isolation of ovine fetal hepatocytes that retain some of the characteristics of the ovine fetal liver while maintained in short-term culture. Supported in part by grants #HD20761, DK 35836, IP30 HD 27827, and HD 00781 from the National Institutes of Health, Bethesda, MD.     

Arachidonic acid metabolism in growth control of A549 human lung adenocarcinoma cells

The role of individual eicosanoids of the arachidonic acid (AA) cascade in the growth control of A549 human lung adenocarcinoma cells has been studied. Cyclooxygenase and lipoxygenase metabolites of [¹⁴C]AA incorporated were actively synthesized in the cultures of tumor cells with full confluence unaccomplished. In such cultures inhibitors of AA metabolism (indomethacin and esculetin) and also a lipoxygenase metabolite of AA, 15-hydroxyeicosatetraenoic acid (15-HETE), significantly suppressed the incorporation of [³H]thymidine and biosynthesis of prostaglandin E₂(PGE₂). Other lipoxygenase metabolites of AA (5-HETE and 12-HETE) had no effect on these parameters. The basic fibroblast growth factor (bFGF) had practically no affect on the growth of A549 cells and the PGE₂ production in cultures with 5% fetal calf serum (FCS); however, in the presence of 0.5% FCS this factor significantly increased the number of tumor cells. The growth-stimulating effect of bFGF was completely abolished by a cyclooxygenase inhibitor indomethacin. The data suggest a key role of PGE₂ in the growth control of A549 cells with an active synthesis of cyclooxygenase and lipoxygenase metabolites of AA, its importance in realization of the mitogenic effect of bFGF, and specific features of 15-HETE as a down-regulator of the PGE₂-dependent proliferation.     

Effect of CO2 on uninfected Sf‐9 cell growth and metabolism

A problem in the mass production of recombinant proteins and biopesticides using insect cell culture is CO₂ accumulation. This research investigated the effect of elevated CO₂ concentration on insect cell growth and metabolism. Spodoptera frugiperda Sf‐9 insect cells were grown at 20% air saturation, 27^°C, and a pH of 6.2. The cells were exposed to a constant CO₂ concentration by purging the medium with CO₂ and the headspace with air. The population doubling time (PDT) of Sf‐9 cells increased with increasing CO₂ concentration. Specifically, the PDT for 0‐37, 73, 147, 183, and 220 mm Hg CO₂ concentrations were 23.2 ± 6.7, 32.4 ± 7.2, 38.1 ± 13.3, 42.9 ± 5.4, and 69.3 ± 35.9 h (n = 3 or 4, 95% confidence level), respectively. The viability of cells in all experiments was above 90%, i.e., while increased CO₂ concentrations inhibited cell growth, it did not affect cell viability. The osmolality for all bioreactor experiments was observed to be 300–360 mOsm/kg, a range that is known to have a negligible effect on insect cell culture. Elevated CO₂ concentration did not significantly alter the cell specific glucose consumption rate (2.5–3.2 × 10^?17 mol/cell s), but slightly increased the specific lactate production rate from ?3.0 × 10^?19 to 10.2 × 10^?19 mol/cell s. Oxidative stress did not contribute to CO₂ inhibition in uninfected Sf‐9 cells as no significant increase in the levels of lipid hydroperoxide and protein carbonyl concentrations was discovered at elevated CO₂ concentration. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:465–469, 2016     

The effect of light on IAA metabolism in different parts of maize seedlings in correlation with their growth

The inhibitory effect of light on the growth of plantscorrelates with a decrease of free IAA content in their tissues andmight be mediated through changes of IAA metabolism. In different partsof Zea mays L. seedlings (roots, mesocotyls and coleoptiles)that respond to light with a different growth rate, the effect of lighton the formation of IAA metabolites was examined in feeding experimentswith ¹⁴C-IAA. In all tissues, IAA was taken up andmetabolised mainly into six compounds, four of them were tentativelyidentified as IAA-1-O-glucose (IAGlc), IAA-myo-inositol, indoleacetamide and IAA-aspartate (IAAsp). IAA was metabolised most slowly inthe roots. In coleoptiles and mesocotyls, IAGlc was the most abundantmetabolite, except for mesocotyls in the light. In roots, a relativelylarge amount of IAA was also metabolised into IAAsp. Light stimulatedthe rate of IAA metabolism in all tissues, but its effect on theconversion of IAA was exceptionally high in mesocotyls. In mesocotyltissue the conversion into IAAsp was greatly stimulated by light.Conversely, the content of IAGlc in mesocotyls was decreased by light.Since light inhibited mesocotyl growth significantly and specifically,it is possible that the high capacity of mesocotyls to synthesise IAAspin the light may have caused a depletion of free IAA, which then led toan inhibition of growth. In mesocotyls from the light-grown plants IAAconjugated into IAGlc was probably used for IAAspbiosynthesis.