Adventitious Bud Formation from Bulb-scale Explants of Lilium speciosum Thunb. in vitro Effects of aminoethoxyvinyl-glycine, 1-aminocyclopropane-1-carboxylic acid,and ethylene

Several representatives of marine brown macroalgae (Phaeophyceae) including Fucus serratus L., Fucus spiralis L. and Fucus vesiculosus L. as well as Laminaria digitata (Huds.) Lamour., Laminaria hyperborea (Gunn.) Foslie and Laminaria saccharina (L.) Lamour. were investigated with particular regard to features of biosynthesis of the storage product mannitol. The respective catalytic system involved in the last step of mannitol formation, mannitol-1-phosphate dehydrogenase, appears to be a cytoplasmic enzyme as may be judged from the degree of correlation with the chloroplast key enzyme ribulose-1,5-bisphosphate carboxylase in different tissues of Laminaria digitata and Laminaria saccharina. Activity of mannitol-1-phosphate dehydrogenase in vitro is not affected by mannitol-l-phosphate or free mannitol, suggesting that mannitol biosynthesis in vivo) is mainly controlled by the environment and/or developmental stage. Certain inorganic ions such as NO₃^- (including K⁺) exert a strong influence on the activity of mannitol1-phosphate dehydrogenase thus suggesting that the intracellular pools of stored NO₃^- and mannitol are confined to spatially separated cellular compartments.     

Segment- and nest-specific determination in the histoblasts of the housefly

Summary Using a novel grafting procedure for histoblasts, we have transplanted the fifth dorsal or ventral histoblast nests to heterotopic positions in the abdomen of the prepupa of the housefly to find out how rigid are these imaginai cells in their commitment to form their respective segmental pattern. Our results clearly show that these histoblasts survive in their new positions and form patterns according to their original determined state.     

Metabolic links between the biosynthesis of pyrrolizidine alkaloids and polyamines in root cultures of Senecio vulgaris

Isotope feeding and inhibitor experiments were performed in order to elucidate the pathway common to polyamine and alkaloid biosynthesis in root cultures of Senecio vulgaris L. -Difluoromethylarginine, a specific inhibitor of arginine decarboxylase, prevented completely the incorporation of radioactivity from [¹⁴C]arginine and [¹⁴C]ornithine into spermidine and the pyrrolizidine alkaloid senecionine N-oxide. In contrast, -difluoromethylornithine, a specific ornithine-decarboxylase inhibitor, had no effect on the flow of radioactivity from labelled ornithine and arginine into polyamines and alkaloids. Thus, putrescine, the common precursor of polyamines and pyrrolizidine alkaloids, is exclusively derived via the arginine-agmatine route. Ornithine is rapidly transformed into arginine. Recycling of the guanido moiety of agmatine back to ornithine can be excluded. Putrescine and spermidine were found to be reversibly interconvertable and to excist in a highly dynamic state. In contrast, senecionine N-oxide did not show any turnover but accumulated as a stable metabolic product. In-vivo evidence is presented that the carbon flow from arginine into the polyamine/alkaloid pathway may be controlled by spermidine. The possible importance of the metabolic coupling of pyrrolizidine-alkaloid biosynthesis to polyamine metabolism is discussed.Abbreviations DFMA D,l--difluoromethylarginine - DFMO D,l--difluoromethylornithine - FW fresh weight     

Expression of gooseberry-proximal in the Drosophila developing nervous system responds to cues provided by segment polarity genes

Summary Segment polarity genes define the cell states that are required for proper organization of each metameric unit of the Drosophila embryo. Among these, the gooseberry locus has been shown to be composed of two closely related genes which are expressed in an overlapping single-segment periodicity. We have used specific antibodies raised against the protein product of the gooseberry proximal (gsb-p) gene to determine the spatial distribution of this antigen in wild type embryos, and to monitor the effects of segment polarity mutants on the pattern of the gsb-p protein distribution. We find that the gsb-p protein accumulates beneath each posterior axonal commissure in the progeny of neuroblasts deriving from the epidermal compartments of wingless (wg) and engrailed (en) expression. The results of this analysis support the idea that gsb-p has a specific role in the control of cell fates during neurogenesis, and indicate that en and wg provide critical positional cues to define the domain in which gsbp will be activated. Furthermore, these data suggest that, in order to be expressed in the embryonic CNS, gsb-p may preliminarily require activity of the gooseberry-distal gene in the epidermis. Offprint requests to: S. Côté     

The kinetics of oocyte activation and polar body formation in bovine embryo clones.

The kinetics of polar body formation were examined in parthenogenetically activated, in vitro matured and aged bovine oocytes. Subsequently, the presence or absence of polar body formation was determined in bovine embryo clones. Polar body formation, defined as telophase II, occurred by 1 (13/40, 43%) and 2 h (15/21, 71%) postparthenogenetic activation of metaphase II stage oocytes. Parthenogenetically activated oocytes readily formed pronuclei by 4 h. Some oocytes had chromatin in a highly condensed state at 1, 2, and 4 h postactivation (13/72, 18%). These oocytes often (10/13, 77%) appeared to be "self-enucleated," as the condensed chromatin was found in a membrane-bound extrusion. The phenomenon was most prevalent when oocytes were handled at room temperature (25-27 degrees C). Nuclear transfer procedures were established to bring about synchronous blastomere fusion and oocyte activation conditions. Synchronous conditions were achieved only when oocytes were handled and manipulated at 37-39 degrees C. Embryo clones examined 2 h postfusion did not form a polar body. Conversely, nucleate demi-oocyte controls were at the late telophase II stage of meiosis. The results are discussed in relation to cell cycle effects on bovine nuclear transfer.     

Fate of microinjected genes in preimplantation mouse embryos.

The state of genes microinjected into mouse embryos was followed from the one-cell to the blastocyst stage using the polymerase chain reaction (PCR). Microinjected DNA was detected in all one-, two-, and four-cell injected embryos and in 44% of morula and 26% of blastocysts. Head-to-tail ligation of microinjected genes, a common feature of stably integrated transgene arrays, was detected in all embryos after injection of microinjected genes and occurred irrespective of the structure at the ends of the injected genes. Sensitivity of microinjected DNA to a methylation-dependent restriction endonuclease Dpn I was lost in all embryos by the two-cell stage (24 hr), indicating a change in DNA methylation, independent of transgene integration. Dissociation of blastomeres prior to compaction revealed a mosaic distribution of the microinjected DNA within the embryo and supports the notion that injected genes form a limited number of arrays, which segregate independently until they integrate into the genome or are degraded.     

Methemoglobin Formation from Butylated Hydroxyanisole and Oxyhemoglobin. Comparison with Butylated Hydroxytoluene and P-Hydroxyanisole

The widely used food additives butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT) react with oxyhemoglobin, thereby forming methemoglobin. The reaction rates were measured using visible spectroscopy, and second order rate constants were established for BHA and compared with p-hydroxyanisole. Using ESR we investigated the involvement of free radical reaction intermediates. The expected one-electron oxidation product of BHA and BHT, the phenoxyl radical, could only be detected with pure 3-t-butyl-4-hydroxyanisole and oxyhemoglobin. With the commercial mixture of 2- and 3-t-butyI-4-hydroxyanisole a very strong ESR signal of a secondary free radical species was observed, similar to the one observed earlier with p-hydroxyanisole and dependent on the presence of free thiol groups, so that we assumed the intermediate existence of a perferryl species, the MetHb-H₂O₂ adduct. In a second series of experiments we investigated the reactivity of this postulated intermediate with BHA and BHT, starting with a pure MetHb/H₂O₂-phenol mixture in a stopped-flow apparatus linked to the ESR spectrometer, detecting the expected phenoxyl radicals from BHA and p-hydroxyanisole. Due to the low solubility and decreased reactivity of BHT only traces of the phenoxyl type radical were found together with a high concentration of unreacted perferryl species. The reactivity of BHA, BHT and p-hydroxyanisole with free thiol groups is demonstrated by an increased reaction rate in the presence of the thiol group blocking substance NEM.     

External pH Modifies the Intracellular pH and the Mode of Photosynthetic CO2-Assimilation in Photoautotrophic Cell Suspension Cultures of Chenopodium rubrum L.

Photoautotrophic cell supension cultures of C. rubrum exhibit a C₃-type of photosynthesis. Yet, up to 20% of total CO₂-fixation is directly incorporated into malate and aspartate during short-term photosynthesis. The rate of ¹⁴C-labeling of malate and aspartate was doubled if the pH of the medium of the cells was increased from the normal value of 4.5 to 6.0 or 7.0. In vivo ³¹P-NMR spectroscopy demonstrated that an increase in the external pH from 4.5 to 6.3 increased the cytosolic pH by 0.3 units and the vacuolar pH by about 1.3 units. Possible mechanisms for the effect of extracellular pH on intracellular pH and PEP-carboxylase-dependent carboxylation reactions are discussed.     

微核形成与细胞周期关系的初步研究——Ⅳ.化学诱变剂诱发人淋巴细胞间期各阶段的微核形成

本实验应用具有诱变作用的抗癌药:噻地哌、长春新碱,乙双吗啉等,体内或体外处理诱发人体外周血淋巴细胞微核,通过控制细胞培养时间,放射性自显影及中期细胞阻滞等方法,定量地分析了细胞间期各阶段的微核率(MNF)。本组实验结果表明,间期各阶段均可有不同程度的微核形成,其中最多的是G_1期,其次是G_2期和G_0期。S期细胞的MNF较G_1期有极显著的下降,这提示大部分G_1期的微核细胞不能进入S期,使细胞增殖中止,这可能是抗癌药物杀伤肿瘤细胞的机制之一。     

A new putative cellulose-synthesizing complex ofColeochaete scutata

Summary Cells of the charophycean alga,Coleochaete scutata active in cell wall formation were freeze fractured in the search for cellulose synthesizing complexes (TCs) since this alga is considered to be among the most advanced and a progenitor to land plant evolution. We have found a new TC which consists of two geometrically distinctive particle complexes complementary to one another in the plasma membrane and occasionally associated with microfibril impressions. In the E-fracture face is found a cluster of 8–50 closely packed particles, each with a diameter of 5–17 nm. Most of these particles are confined within an 80 nm circle. In the P-fracture face is found an 8-fold symmetrical arrangement of 10 nm particles circumferentially arranged around a 28 nm central particle. The TCs ofC. scutata are quite distinctive from the rosette/globule TCs of land plants. The 5.5×3.1 nm microfibril inC. scutata is also distinctive from the 3.5×3.5 nm microfibril typical of land plants. The phylogenetic implications of this unique TC in land plant evolution are discussed.