Elicitor-induced metabolic changes in cell cultures of chickpea (Cicer arietinum L.) cultivars resistant and susceptible to Ascochyta rabiei

Cell-suspension cultures of two chickpea (Cicer arietinum L.) cultivars, resistant (ILC 3279) and susceptible (ILC 1929) to the fungus Ascochyta rabiei (Pass.) Lab., showed differential accumulation of the phytoalexins medicarpin and maackiain, and transient induction of related enzyme activities after application of an A. rabiei-derived elicitor. The chalcone-synthase (CHS) activity (EC 2.3.1.74) which is involved in the first part of phytoalexin biosynthesis exhibited a maximum 8–12 h after elicitation in the cells of both cultivars. Concomitant with the fivefold-higher phytoalexin accumulation, CHS activity increased twofold in the cells of the resistant cultivar. The maximum of the elicitor-induced CHS-mRNA activity was determined 4 h after onset of induction in the cultures of both cultivars, although in cells of cultivar ILC 3279 this mRNA activity was induced at a level twofold higher than that in cells of the susceptible race ILC 1929. Investigations of CHS isoenzymes by two-dimensional gel electrophoresis of immunoprecipitated in-vitro-translated protein indicated the presence of five proteins. In the cells of both cultivars only two of the isoenzymes were induced after elicitor treatment. Analysis of the total in-vitro-translated proteins by two-dimensional gel electrophoresis showed that the constitutively expressed patterns of mRNA activities in the cell cultures of the two cultivars were identical. After elicitation, considerably more translatable mRNAs were induced in the cells of cultivar ILC 3279. The few induced proteins, and their respective mRNA activities, which could be detected in the cells of the susceptible cultivar, all existed in the cells of the resistant cultivar, too. One highly induced protein (M_r 18 kDa) found in the cells of cultivar ILC 3279 reached its maximum mRNA activity 6 h after elicitor application. The amount of this protein was hardly increased in the cells of the susceptible cultivar. This protein appears to be excreted from the cells into the growth medium.Abbreviations CHS chalcone synthase - IEF isoelectric focussing - ILC international legume chickpea - PR-protein pathogenesis-related protein - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis Financial support by Deutsche Forschungsgemeinschaft and Fonds der Chemischen Industrie is gratefully acknowledged. The authors thank Dr. K. Hahlbrock (Max-Planck-Institut für Züchtungsforschung, Köln, FRG) for provision of antisera and the International Centre for Agricultural Research in the Dry Areas (Aleppo, Syria) for plant material.     

Different influences of cytochalasin B on the activation of human neurophils settled onto petri dishes displayed by simultaneously detected native and luminol-dependent luminescence

Cytochalasin B (CB) is known to interfere reversibly with the cytoplasmic contractile filamental network of mammalian cells. The role of the microfilament system in the mechanism of the reactive oxygen intermediates release of polymorphonuclear leukocytes (PMNL) was studied for different kinds of stimuli. PMNL from fresh human blood were treated with CB and stimulated by adherence on plastic surfaces, by opsonized zymosan, by phorbol myristate acetate and by N-formylmethionyl-phenylalaline. The production of reactive oxygen species were monitored by simultaneous detection of native, luminol-independent, luminescence (NL) and luminol-dependent luminescence (LDL) using a method of spectral discrimination. Different influences of CB on NL with respect to LDL as well stimuli-dependent influences of CB on the luminescence response of PMNL were observed. Especially phagocytosis-associated activation of PMNL was strongly inhibited by CB, whereas LDL was reduced to a much greater extent in comparison with NL. A firm involvement of the microfilament system is indicated, but it depends on the kind of stimulus engaged.     

Langzeit-Erfassung brutbiologischer Parameter beim KleiberSitta europaea in zwei norddeutschen Untersuchungsr?umen

Zusammenfassung Der Zusammenstellung liegen Befunde zu den brutbiologischen Parametern Gelegestärke, Schlüpftermin und Anzahl flügger Junge zugrunde, die von 1970–1995 an rund 1500 Erstbruten des Kleibers in Versuchsgebieten mit künstlichen Nisthöhlen bei Braunschweig und Essen gewonnen wurden. Die Vollgelege-Eizahl beträgt im Durchschnitt 6,8 Eier (nur Daten aus dem Braunschweiger Raum), der durchschnittliche Schlüpftermin fällt auf den 9. Mai (Braunschweig) und 6. Mai (Essen) und die Anzahl flügger Nestlinge liegt in erfolgreichen Bruten im Mittel bei 5,6 (Braunschweig) bzw. 5,2 (Essen). Im Untersuchungszeitraum kam es in beiden Gebietsregionen zu einer signifikanten Verfrühung des Bruttermins. Doch werden wohl erst die Befunde der kommenden Jahre zeigen können, ob die rezente Häufung früher Brutjahre beim Kleiber (diese Arbeit) und bei Kohl- und Blaumeisen (Winkel &Hudde unveröff.) bereits eine Folge des anthropogen bedingten global warming ist oder noch das normale Auf und Ab der Witterung widerspiegelt.

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Long-term changes of breeding parameters of NuthatchesSitta europaea in two study areas of northern Germany

Between 1970 and 1995, nestbox data about clutch size, hatching date and fledging success were collected for a total of around 1500 first broods of Nuthatches in two north German study areas (near Brunswick 52.16 N 10.32 E and near Essen 51.24 N 6.59 E). The average clutch size was 6.8 eggs (Brunswick), the average hatching date 9 May (Brunswick) and 6 May (Essen), and the mean number of fledglings per successful brood 5.6 (Brunswick) and 5.2 (Essen). Over time hatching dates significantly advanced in both areas. Data of further years are needed to clarify if the recent unusual sequence of early breeding years in the Nuthatch is already a consequence of the anthropogenic greenhouse effect or just due to normal variation of weather conditions.

     

Agglutinierte Foraminiferen aus Graptolithen-Schwarzschiefern des Llanvirnium (Ordovizium) von Plettenberg im Sauerland (Nordrhein-Westfalen,NW-Deutschland)

The Plettenberger Bänderschiefer (Ordovician, Lower Llanvirnian, Zone ofDidymograptus bifidus (Hcall) from the type locality at Plettenberg (Sauerland, NW-Germany) yielded a foraminiferal fauna consisting of primitive agglutinated species (Amphitremoida, Bathysiphon,?Raibosammina, ?Thekammina, Thurammina). Forming one of the stratigraphically oldest European foraminiferal assemblages, these taxa were washed out of an unconsolidated graptolite siltstone and represent a pelagic, oxygen-depleted environment.     

Succinate-ethanol fermentation in Clostridium kluyveri: purification and characterisation of 4-hydroxybutyryl-CoA dehydratase/vinylacetyl-CoA Δ3-Δ2-isomerase

Anaerobically prepared cell extracts of Clostridium kluyveri grown on succinate plus ethanol contained high amounts of 4-hydroxybutyryl-CoA dehydratase, which catalyzes the reversible dehydration of 4-hydroxybutyryl-CoA to crotonyl-CoA. The enzyme was purified 12-fold under strictly anaerobic conditions to over 95% homogeneity and had a specific activity of 123 nkat mg^-1. The finding of this dehydratase means that all of the enzymes necessary for fermentation of succinate plus ethanol by C. kluyveri have now been demonstrated to exist in this organism and confirms the proposed pathway involving a reduction of succinate via 4-hydroxybutyrate to butyrate. Interestingly, the enzyme is almost identical to the previously isolated 4-hydroxybutyryl-CoA dehydratase from Clostridium aminobutyricum. The dehydratase was revealed as being a homotetramer (m=59 kDa/subunit), containing 2±0.2 mol FAD, 13.6±0.8 mol Fe and 10.8±1.2 mol inorganic sulfur. The enzyme was irreversibly inactivated after exposure to air. Reduction by sodium dithionite also yielded an inactive enzyme which could be reactivated, however, up to 84% by oxidation with potassium hexacyanoferrate(III). The enzyme possesses an intrinsic vinylacetyl-CoA isomerase activity which was also found in 4-hydroxybutyryl-CoA dehydratase from C. aminobutyricum. Moreover, the N-terminal sequences of the dehydratases from both organisms were found to be 63% identical.     

Glycosylation in cardenolide biosynthesis

The glycosylation and deglycosylation of cardiac glycosides was investigated using cell suspension cultures and shoot cultures, both established from Digitalis lanata EHRH. plants, as well as isolated enzymes. Shoots were capable of glucosylating digitoxigenin, evatromonoside, digiproside, glucodigitoxigenin and digitoxin. Suspension cultured Digitalis cells glucosylated all the substrates mentioned but digiproside, whereas the UDP-glucosedependent cardinolide glucosyltransferase isolated from that source did not accept digitoxigenin and digiproside as substrates. It is concluded that at least three different glucosyltransferases are involved in cardiac glycoside formation in Digitalis. Similar experiments carried out with glucosylated cardenolides which were administered to cultured cells, shoots and a cardenolide -glucosidase isolated from young leaves revealed that at least two different glucosidases occur in Digitalis lanata, albeit in different tissues or during different phases of development. The biotransformation of glucoevatromonoside was investigated using unlabelled compound and [¹⁴C-glucose]-glucoevatromonoside synthesized enzymatically. After 7 d of incubation almost no radioactivity could be recovered from the cardenolide fraction, indicating that the terminal glucose of glucoevatromonoside was now incorporated into volatile, hydrophilic and insoluble compounds. Since, on the other hand, large amounts of cardenolides were found in the experiments with unlabelled glucoevatromonoside it is assumed that steady state or pool size regulation is achieved by the coordinated action of a cardenolide glucosidase and a glucosyltransferase.Abbreviations Acdox D-acetyldigitoxose - dgen digoxigenin - dox D-digitoxose - dten digitoxigenin - dtl D-digitalose - fuc D-fucose - gten gitoxigenin - qun D-quinovose - CGH cardenolide 16-O-glucohydrolase - DFT UDP-fucose:digitoxigenin 3-O-fucosyltransferase - DGT UDP-glucose:Digitoxin 16-O-glucosyltransferase - DQT UDP-quinovose:digitoxigenin 3-O-quinovosyltransferase     

Folate responsiveness during growth and development of Dictyostelium: separate but related pathways control chemotaxis and gene regulation

Folate-controtled gene expression and chemotaxis have been examined in Dictyostelium wild-type and mutant strains. We show that regulation of the discoidin genes is sensitive to foiate in growing ceiis as weli as in suspension development. The signal is transferred via the N¹⁰-methylfoiate-sensitive folate receptor sites, which also appear to confer the chemotactic response. The strain HG5145 has previously been isolated as a mutant that does not display chemotactic movement towards folate. Nevertheless, these cells are fully functional in foiate-mediated downregulation of discoidin I expression. The strain ga 93 has been isolated as an overproducer mutant of the cyclic nucleotide phosphodiesterase inhibitor. Simultaneously, these cells fail to downregulate discoidin I in response to folate but are fully functional in folate chemotaxis. Therefore we conclude that the pathways for chemotaxis and for gene regulation diverge downstream of a common receptor type.