Specificity of rabbit antisera against the rough lipopolysaccharide of Salmonella minnesota strain R7 (chemotype Rd1P−)

Abstract Rabbit polyclonal antibodies against the lipopolysaccharide (LPS) of the Rd₁P⁻ mutant strain R7 of Salmonella minnesota were serologically characterized using R7 LPS, dephosphorylated LPS, deacylated LPS, deacylated, dephosphorylated and reduced LPS, and synthetic partial structures. The latter comprised partial structures of the core region of Rd₁P⁻ LPS bound to the β 1 → 6-linked glucosamine disaccharide with two amide-linked 3-hydroxytetradecanoic acid residues or artificial glycoconjugates comprised of the synthetic oligosaccharides coupled to bovine serum albumin. Using a passive hemolysis and an enzyme immunoassay, absorption and inhibition experiments, the antibody specificities present could be determined. One group of antibodies required components of the core region and the phosphorylated glucosamine disaccharide of the lipid A moiety for binding. The majority of phosphate-independent antibodies was directed against the trisaccharide l -glycero-α- d -manno-heptopyranose(1 → 3)- l -glycero-α- d -manno-heptopyranose(1 → 5)3-deoxy- d -manno-octulosonic acid. Antibodies against the 1 → 3- and 1 → 7-linked heptose disaccharides and against a single heptose were also detected, however, with low titers. No antibodies were found which required the presence of fatty acids.     

Über die elektrische Kapazität der Zellmembran und die Leitfähigkeit des Zytoplasmas von Ehrlich-Aszitestumorzellen

Zusammenfassung In der vorliegenden Arbeit wird die elektrische Leitfähigkeit und die Membrankapazität von Ehrlich-Aszitestumorzellen gemessen. Die Membrankapazität beträgt 1,7±0,3 Farad/cm². Die spezifische Leitfähigkeit des Zytoplasmas beträgt 0,013 [ ^–1 cm^–1]±12%. Der spezielle Verlauf der Dispersion der Dielektrizitätskonstanten und der Leitfähigkeit der Zellsuspension zeigt, daß ein sehr breites Spektrum von Relaxationszeiten vorliegt, das nicht durch die Größenverteilung der Zellen allein erklärt werden kann. Die Spektralverteilung der Relaxationszeiten hat die FormH(T)=const. Die spezifische Leitfähigkeit des Protoplasmas kann in erster Näherung durch die elektrische Beweglichkeit der Elektrolytionen in einer etwa 15%igen Proteinlösung erklärt werden.Für die Mitarbeit bei den Versuchen möchte ich Frau H.Valetas meinen Dank aussprechen.Herrn Prof. Dr. Dr. h. c. Dr. h. c.Boris Rajewsky zum 70. Geburtstag gewidmet.     

Entwicklungsgeschichtliche untersuchungen an einer Blattmutante vonHyoscyamus niger L.

Summary The leaf shape of the mutantfiliformis (fil) ofHyoscyamus niger L. is strongly modified by external factors (like nutrition and light) as well as by the height of insertion. The name filiformis refers to thread-like leaves which always occur in the inflorencence; they may also be formed in the vegetative region, especially under short day conditions. Other leaves may have a small rhombic blade or a larger blade with irregular edges and deep incisions. Even pinnate leaves have been found. In contrast to the leaves of normalHyoscyamus, all mutant leaves (hypsophylls included) have a stalk-like basal portion that seems to be homologous to the basal part of the normal blade. This mutant is caused by one recessive factor which is linked neither toann nor topall.the submarginal initials of the normalHyoscyamus blade were always found dividing according to the periclinal-anticlinal type, while in the mutant the activity of the submarginal initials frequently resulted in a primarily biseriate mesophyll (so-called double-edged segmentation).This is apparently the first time that gene control of the mode of submarginal blade growth has been observed. Further differences between mutant and normalHyoscyamus concern the venation, the lengths of palisade cells and of stomata guard cells, the frequency of stomata per mm², and the thickness of the blade.

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Electron Microscopy of Peripheral Nerves and Neuromuscular Junctions in the Wasp Leg

The peripheral nerve branch innervating the femoral muscles of the common yellow jacket (Vespula carolina) has been found to possess a thick lemnoblast basement membrane and a complex mesaxon. The term "tunicated nerve" is proposed to designate the type of peripheral nerve in which one or several axons are loosely mantled by meandering, cytoplasm-enclosing membranes of the lemnoblast. The peripheral axon courses longitudinally in a groove in the muscle fiber between the plasma membrane of the muscle fiber and a cap formed by lemnoblast and tracheoblast. The junction is characterized by apposition of plasma membranes of axon and muscle fiber, abundant mitochondria, and synaptic vesicles in the axon, and aggregates of "aposynaptic granules" plus mitochondria and endoplasmic reticulum on the muscle side of the synapse. Unlike the vertebrate striated muscle fiber, no complex infolding of the synapsing plasma membrane of the muscle fiber occurs. The "connecting tissue" of the insect is formed by tracheoblasts, their basement membranes, and the basement membranes of other cells. Further mechanical support is given by the ramifying tracheoles. The physiologic roles of the specialized structures are considered.     

Benzothiadiazole induces disease resistance in Arabidopsis by activation of the systemic acquired resistance signal transduction pathway

Benzothiadiazole (BTH) is a novel chemical activator of disease resistance in tobacco, wheat and other important agricultural plants. In this report, it is shown that BTH works by activating SAR in Arabidopsis thaliana. BTH-treated plants were resistant to infection by turnip crinkle virus, Pseudomonas syringae pv ‘tomato’ DC3000 and Peronospora parasitica. Chemical treatment induced accumulation of mRNAs from the SAR-associated genes, PR-1, PR-2 and PR-5. BTH treatment induced both PR-1 mRNA accumulation and resistance against P. parasitica in the ethylene response mutants, etr1 and ein2, and in the methyl jasmonate-insensitive mutant, jar1, suggesting that BTH action is independent of these plant hormones. BTH treatment also induced both PR-1 mRNA accumulation and P. parasitica resistance in transgenic Arabidopsis plants expressing the nahG gene, suggesting that BTH action does not require salicylic acid accumulation. However, because BTH-treatment failed to induce either PR-1 mRNA accumulation or P. parasitica resistance in the non-inducible immunity mutant, nim1, it appears that BTH activates the SAR signal transduction pathway.     

The leaf peroxisomal form (MFP IV) of multifunctional protein functioning in fatty-acid β-oxidation

We have purified for the first time from green leaves a multifunctional protein (MFP) involved in fatty acid -oxidation. The protein, designated MFP IV, was extracted from green leaves of three-week-old cucumber (Cucumis sativus L.) plants. Chromatography on cation exchanger, separation on hydroxylapatite, and fast-protein liquid chromatography on Phenylsuperose led to a more than 7000-fold purification and to the isolation of an apparently homogeneous 80-kDa monomeric protein. This protein is immunologically related to the glyoxysomal MFP II, as evidenced by immunodecoration with antiserum raised against MFP II. Comparison of molecular masses of all MFPs presently known revealed that the MFP prepared from green leaves (MFP IV) is distinct from MFP II (76.5 kDa) and MFP I (74 kDa) from dark-grown cotyledons. By including other properties in this comparison, we demonstrated that MFP IV can also be distinguished from the glyoxysomal MFP III (81 kDa) and the bacterially expressed MFP-a (80 kDa). Moreover, MFP IV is a constituent of leaf peroxisomes and contains the activities of 2-enoyl-CoA hydratase (EC 4.2.1.17),l-3-hydroxyacyl-CoA dehydrogenase (EC 1.1.1.35) and 3-hydroxyacyl-CoA epimerase.Abbreviation MFP multifunctional protein This work was supported by grants from the Deutsche Forschungsgemeinschaft and the Fonds der Chemischen Industrie.     

ent-Kaurene biosynthesis in a cell-free system from wheat (Triticum aestivum L.) seedlings and the localisation of ent-kaurene synthetase in plastids of three species

A cell-free system capable of converting [¹⁴C]geranylgeranyl diphosphate to ent-[¹⁴C]kaurene and to an unidentified acid-hydrolysable compound was obtained from the basal portions of 5-d-old shoots of wheat seedlings (Triticum aestivum L.). By means of marker enzyme activities, the synthesis of ent-kaurene and the unknown compound could be quantitatively assigned to a plastid fraction obtained by Percoll-gradient centrifugation of the homogenate. The enzyme activities were located within the plastids, probably in the stroma, because they withstood trypsin treatment of the intact plastids, and the plastids had to be broken to release the activity, which was then obtained in soluble form. Plastid membranes had no activity. Plastid stroma preparations obtained from pea (Pisum sativum L.) shoot tips and pumpkin (Cucurbita maxima L.) endosperm also yielded ent-kaurene synthetase activity, but did not form the unknown compound. The exact nature of the active plastids was not ascertained, but the use of methods for proplastid isolation was essential for full activity, and the active tissues are all known to contain high proportions of proplastids, developing chloroplasts or leucoplasts. We therefore believe that ent-kaurene synthesis may be limited to these categories. Mature chloroplasts from the wheat leaves did not contain ent-kaurene synthetase activity and did not yield the unknown component. Incorporation of [¹⁴C]geranylgeranyl diphosphate into ent-[¹⁴C]kaurene and the unknown component was assayed by high-performance liquid chromatography with on-line radiocounting. ent-[¹⁴C]Kaurene was identified by Kovats retention index and full mass spectra obtained by combined gas chromatography-mass spectrometry. The unknown component was first believed to be copalyl diphosphate, because it yielded a compound on acid hydrolysis, which migrated like copalol on high-performance liquid chromatography and gave a mass spectrum very similar to that of authentic copalol. However, differences in the mass spectrum and in retention time on capillary gas chromatography excluded identity with copalol. Furthermore, the unhydrolysed compound was not converted to ent-kaurene by a cell-free system from C. maxima endosperm as copalyl diphosphate would have been.Abbreviations ADH alcohol dehydrogenase - AMO 1618 2isopropyl-4-(trimethylammoniumchloride)-5-methylphenyl piperi-dine-1-carboxylate - BSA bovine serum albumin - DTT dithioth-reitol - GA_n gibberellin A_n - GAPDH NADP⁺-glyceraldehyde 3-phosphate dehydrogenase - GC-MS combined gas chromatography-mass spectrometry - GGPP all trans-isomer of geranyl-geranyl diphosphate - KS ent-kaurene synthetase - MDH malate dehydrogenase - MAA mevalonate activating activity - SOR shikimate oxidoreductase We thank Mrs. Gudrun Bodtke and Mrs. Dorothee Dasbach for able technical assistance, Prof. L.N. Mander (Australian National University, Canberra, Australia) for ent-[²H₂]kaurene and Dr. Yuji Kamiya (RIKEN, Saitama, Japan) for geranylgeraniol and copalol. The work was supported by the Deutsche Forschungsgemeinschaft.     

Sodium channels in cultured neuroblastoma cells grown in high glucose or l-Fucose

Patch clamp techniques were used to record whole cell and single channel Na⁺ currents from NB41A3 neuroblastoma cells grown in culture. Cells were grown for two weeks in control medium or medium supplemented with 30 mm d-glucose of 30 mm l-fucose.Cells exposed to glucose or l-fucose had smaller whole cell Na⁺ currents than cells grown in unsupplemented medium, consistent with earlier studies (Yorek, Stefani & Wachtel, 1994). Whole cell macroscopic currents showed no change in activation or inactivation kinetics. Single channel current properties and opening probability were also unchanged.The number of [³H]saxitoxin binding sites, and therefore the total number of Na⁺ channels, was not reduced in cells grown in glucose or l-fucose (Yorek et al., 1994). Therefore, we conclude that some of the channels must have been rendered nonfunctional by the conditioning media. The finding that single channel properties are not altered suggests that channels become nonfunctional in an all-or-none manner.This work was supported by Merit Review Awards to M.A. Yorek and R.E. Wachtel from the Department of Veterans Affairs and by National Institutes of Health grant DK45453 to M.A. Yorek.