Comparative effects of adenine analogs upon metabolic cooperation between Chinese hamster cells with different levels of adenine phosphoribosyltransferase activity

Colony formation by variant Chinese hamster cells highly resistant to adenine analogs and deficient in adenine phosphoribosyltransferase (APRT) activity was measured after co-cultivation with APRT⁺, CHO-K1 cells in medium containing one of three different adenine analogs. Depending upon the density of APRT⁺ cells and the specific adenine analog, large differences in the recovery of APRT^? colonies were observed. The particular adenine analog and APRT⁺ cell density were more significant factors in the recovery of APRT^? colonies than the concentration of the analog or the level of APRT activity. The number of wild-type cells (CHO-K1) required to inhibit formation of APRT^? colonies by 50% (mean lethal density; MLD₅₀) with 65 μg/ml 8-aza-adenine (AzA) as the selective drug was 8.0 × 10⁵ cells/100 mm dish (1.5 × 10⁴/cm²). With 100 μg/ml 2,6-diaminopurine (DAP) the MLD₅₀ for CHO-K1 was 4.0 × 10⁵ cells/100 mm dish (7.3 × 10³/cm²). The MLD₅₀ for CHO-K1 when the DAP concentration was decreased to 50 μg/ml was only slightly higher, 5 × 10⁵ cells/100 mm dish (9.1 × 10³/cm²). The most toxic effect was observed with 2-fluoroadenine (FA). The MLD₅₀ for CHO-K1 in 2 μg/ml FA was 4.5 × 10⁴ cells/100 mm dish (8.2 × 10²/cm²), a cell density which permits minimal direct contact between APRT⁺ and APRT^? cells. The toxic effects of FA on individually resistant, APRT^? cells were found to be mediated by metabolites released into the medium by dying APRT⁺ cells. This metabolite toxicity to APRT^? cells was also demonstrated in mixtures with cells having only 8% of wild-type APRT activity. The MLD₅₀ for these APRT⁺ (8%) cells in 2 μg/ml FA was 7.5 × 10⁴ cells/100 dish (1.4 × 10³/cm²), a small difference from the MLD₅₀ for cells with wild-type levels of APRT activity. The differences in the recovery of APRT^? colonies from mixtures with APRT⁺ cells in these three adenine analogs are critical to the design of procedures for the selection of APRT^? cells from populations of APRT⁺ cells and emphasize the importance of establishing the parameters of metabolic cooperation, not only in terms of cell density but also with regard to the particular selective agent, in any experiment designed to determine precise mutation rates or to test putative mutagens upon mammalian cells in culture.     

大刍草稀有等位基因促进玉米密植高产

密植是提高作物单位面积产量、促进粮食增产的重要途径之一。叶夹角是影响玉米(Zea mays)密植的关键因子。中国农业大学田丰课题组最近克隆了2个调控玉米叶夹角的数量性状位点(QTL)——UPA1和UPA2, 揭示了这2个位点的功能基因(brd1和ZmRAVL1)通过油菜素内酯(BR)信号通路调控叶夹角。UPA2位于ZmRAVL1上游9.5 kb, 可与DRL1蛋白结合。另一个影响玉米叶夹角的蛋白LG1可以激活ZmRAVL1的表达; DRL1蛋白与LG1蛋白直接互作抑制LG1对ZmRAVL1的激活表达。玉米祖先种大刍草(teosinte)的UPA2位点序列与DRL1蛋白结合能力更强, 导致大刍草ZmRAVL1的表达受到更强的抑制, 下调表达的ZmRAVL1进一步使下游基因brd1的表达下调, 进而降低叶环区的内源BR水平, 导致叶夹角变小。将大刍草的UPA2等位基因导入到玉米中或对玉米中ZmRAVL1进行基因编辑, 在密植条件下均可显著提高玉米产量。上述发现为高产玉米品种的分子育种改良提供了重要理论基础和基因资源。     

Differentiation under the control of insulin of rat preadipocytes in primary culture: Isolation of homogeneous cellular fractions by gradient centrifugation

Using a density gradient medium (Percoll) we succeeded in isolating homogeneous cell populations from the stromal-vascular fraction of the inguinal tissue of 3-day-old rats. In primary culture, in medium 199 supplemented with 10% fetal calf serum and 5.5 mM glucose, almost complete differentiation (90%) of these fractions was obtained for the first time in presence of a physiological concentration of insulin (10^?9 M). During the adipose conversion, insulin markedly enhanced the activities of glycerol-3-phosphate dehydrogenase and acid:CoA ligase. When VLDL and heparin were added with insulin to the medium, this effect was not potentiated. On the contrary, VLDL and heparin in presence of insulin increased the triglyceride content of the cells. With VLDL and heparin only, the biochemical and morphological characteristics of the cells were very similar to those observed in control culture. The heavier fraction was morphologically heterogeneous and did not undergo the adipose conversion to the same extent as the two lighter fractions. It was concluded that this model could be helpful in studying the proliferation and the differentiation of preadipocytes at an early stage of development.     

Characterization of a variant of carcinoembryonic antigen (CEA) using monoclonal and polyclonal anti-CEA antibodies

A variant of the carcinoembryonic antigen (CEA) with lower molecular weight than a CEA reference preparation has been separated from CEA. Using a polyclonal, spleen absorbed anti-CEA antiserum, the variant crossreacts with reference CEA in immunodiffusion. The CEA-activity of the variant has been demonstrated using an enzyme-immunoassay with monoclonal CEA specific antibodies. There is sufficient immunological evidence that this variant is a distinct antigen different from the crossreactive antigens described so far. The reactivity of the polyclonal anti-CEA antiserum with the CEA variant was abolished by absorption against the immobilized variant.     

Composition and seasonal fluctuations of the ichthyoplankton community in a shallow tidal channel of Cadiz Bay (S.W. Spain)

The composition and seasonal variation of the ichthyoplankton community in a shallow tidal channel in Cadiz Bay was studied for 5 years (1984 1988), taking fortnightly samples with a conical tide-strained net (0.5 mm mesh). Thirty-nine species belonging to 21 families of teleosts were caught. Four species ( Pomutoschistus microps Kroyer, Atherina hoyeri Risso, Liza rumuda Risso and Lizu a lira Ia Risso) totalled more than 90% of the annual ichthyoplankton abundance. Three species, assumed to be permanent residents, dominated the number offish (837%), and were represented by all their developing stages; 26 species (66.7%) were temporary residents, contributing 17.2% to the total abundance and were represented only by early developing stages; the remaining 10 species were classified as rare. Monthly numbers of species and individuals showed seasonal variation with a main annual peak in spring and in late spring-early summer, respectively. The diversity (H') and evenness (S) indices of this community were low (0.13 to 1.83 and 0.03 to 0.71. respectively), with higher values in winter and late summer, and both were significantly correlated with the log-transformed density of temporary residents (r = 0.43 and 0.41, respectively; P<0.0l) and permanent residents (r=−0.75 and −0.79, respectively; P<0.01). Multivariate classification analysis of samples revealed that similar cyclical changes were taking place in the structure of this ichthyoplankton community every year; three periods could be differentiated: (1) a cool period of low fish density but relatively high diversity; (2) moderate temperature and salinity with high numbers of species and density but low diversity and (3) a warm period of moderate diversity and fish density.     

Influence of phenolglucosides and trichome density on the distribution of insects herbivores on willows

The effects of both trichome density and phenolglucoside content of leaves of 76 willow hybrids (Salix alba x fragilis) were measured to estimate their influence on the distribution of Phratora vitellinae (L.), Plagiodera versicolora Baly (Coleoptera: Chrysomelidae) and Pontania proxima (Lepeletier 1823) (Hymenoptera: Tenthredinidae) in a nursery at Gramont, Belgium.The willows showed differences in their phenolglucoside content and pilosity of leaves and are classified on these basis into four groups by a clustering method. Correlations and multiple regressions showed that these chemical and physical characteristics are good predictors of the abundance of insects. First, the abundance of larvae of Ph. vitellinae, adults of Pl. versicolora and galls of P. proxima is correlated positively with a high phenolglucoside content and a low pilosity of the leaves. Secondly, the distribution of adults of Ph. vitellinae and of larvae of Pl. versicolora is influenced by neither the chemical nor the physical leaves characteristics studied.     

Population structure,growth and fecundity of perch (Perca fluviatilis L.) in an acidified river system in Southern Norway

Population structure, density and reproductive potential of perch (Perca fluviatilis L.) from 4 acidified lakes in a river system in Southern Norway were investigated. The upper 3 lakes were most affected by acidification and the number of perch caught per unit effort indicate three-fold increase in density from the upper Gjerstadvann to the lower Brøbørvann. Low density of perch in the upper lakes is explained by; 1) Abnormal mortality occurring in episodes due to acidification, probably caused by aluminium toxicity at pH = 5.2–5.4. 2). Acid water (pH ≤ 5.0) during spawning and development of eggs and larvae, resulting in recruitment failure. Juvenile mortality of perch may depend on the density of the parental stock. In the 3 upper lakes, the density of adult perch probably was too low to produce significant juvenile mortality, and in these lakes the recruitment probably depended more on the May water quality. Improved water quality increases egg hatching and survival and also benefits zooplankton production, the main food for perch during the first summer. Low population density has reduced competition for food and thus improved growth of perch in the 3 upper lakes compared to the lower lake. The perch in Brøbørvann mature at higher age and have lower individual fecundity than perch in Gjerstadvann, an effect of both slower growth and lower length specific fecundity in Brøbørvann. In Gjerstadvann, the perch therefore have a higher reproductive potential relative to the stock density, and may, when the water quality is good enough, give rise to strong year-classes and more frequent year-class fluctuations than perch in the less acid Brøbørvann.     

The nematode fauna of a temperate Australian mangrove mudflat; its population density,diversity and distribution

The nematode fauna of an estuarine mangrove Avicennia marina mudflat in Southeastern Australia has been intensively studied. About 85% of the nematodes occur in the top cm of soft mud, but 5–7 species inhabit the deeper anoxic mud down to 10 cm, both at low and high tide. One square metre was intensively sampled from four zones with different nematode faunas. At the low tide zone 58% of the nematodes were epistrate feeders, including many diatom-feeders, but in the mangrove zone selective microbial feeders made up over 60% of the population, while between high water neap and high water spring, above the mangrove zone, omnivore/predators and plant root feeding nematodes increased in relative importance. Random replicate cores reliably sampled species occurrence, but gave a high variance in density estimates. Replicate aliquots from homogenised mud gave lower density variance. Nematode densities (maximum 5 × 10⁶ m^-2) were not as high as have been reported from non-mangrove estuaries in other countries, but were within the range found in mangroves elsewhere in Australia. Margalef Species Richness values ranged from 1.7 to 3.89, which is similar to values found in other mangroves mudflats in Australia. Nematode biomass ranged from 888 mg dry weight m² (383 mg C m^-2) at the low tide zone to 19 mg dry weight m^-2 (8 mg C m^-2) at the upper tide level.     

Effect of salinity and pH on cobalt biosorption by the estuarine microalgaChlorella saliva

Summary Accumulation of cobalt (⁶⁰Co) by the estuarine microalgaChlorella salina has been characterized. At cobalt concentrations ranging over 3.125–100 M, a significant amount of cobalt was bound within 1 min. This was metabolism-independent and unaffected by incubation in light or dark conditions. This initial rapid phase of biosorption was followed by a slower phase of uptake which was apparently active and inhibited by incubation in the dark, or by the uncoupler dinitrophenol and the respiratory and photosynthetic inhibitor potassium cyanide in the light. For cells suspended in 10 mM Taps pH 8, cobalt biosorption followed a Freundlich adsorption isotherm. However, in the presence of 0.5 M NaCl, biosorption deviated from the Freundlich model because of competition by Na⁺. Cobalt biosorption was decreased by increasing concentrations of Na⁺, decreasing pH and the presence of Cs⁺, Li⁺, Rb⁺, Zn²⁺. Mn²⁺ and Sr²⁺ (added as chlorides). This was a result of competition between Co²⁺ and the other cations, including H⁺, for available binding sites on the cell wall and was confirmed by increased desorption of cobalt by solutions of low pH or high salinity. Increasing cell density resulted in increased removal of cobalt from solution but decreased the specific amount of cobalt taken up by the cells.