Quantitation of gibberellins and the metabolism of [3H]gibberellin A1 during somatic embryogenesis in carrot and anise cell cultures

In a carrot (Daucus carota L.) cell line lacking the ability to undergo somatic embryogenasis, and in carrot and anise (Pimpinella anisum L.) cell lines in which embryogenesis could be regulated by presence or absence of 2,4-dichlorophen-oxyacetic acid (2,4-D), in the medium (+2,4-D=no embryogenesis,-2,4-D=embryo differentiation and development), the levels of endogenous gibberellin(s) (GA) were determined by the dwarfrice bioassay, and the metabolism of [³H]GA₁ was followed. Embryos harvested after 14 d of subculture in-2,4-D had low levels (0.2–0.3 g g^-1 dry weight) of polar GA (e.g. GA₁-like), but much (3–22 times) higher levels of less-polar GA (GA_4/7-like); GA₁, GA₄ and GA₇ are native to these cultures. Conversely, the undifferentiated cells in a non-embryogenic strain, and proembryos of an embryogenic strain (+2,4-D) showed very high levels of polar GA (2.9–4.4 g g^-1), and somewhat reduced levels of less-polar GA. Cultures of anise undergoing somatic embryo development (-2,4-D) metabolized [³H]GA₁ very quickly, whereas proembryo cultures of anise (+2,4-D) metabolized [³H]GA₁ slowly. The major metabolites of [³H]GA₁ in anise were tentatively identified as GA₈-glucoside (24%), GA₈ (15%), GA₁-glucoside (8%) and the ¹⁽¹⁰⁾GA₁-counterpart (2%). Thus, high levels of a GA₁-like substance and a reduced ability to metabolize GA₁ are correlated with the absence of embryo development, while lowered levels of GA₁-like substance and a rapid metabolism of GA₁ into GA₈ and GA-conjugates are correlated with continued embryo development. Exogenous application of GA₃ is known to reduce somatic embryogenesis in carrot cultures; GA₄ was found to have the same effect in anise cultures. Thus, a role (albeit negative) in somatic embryogenesis for a polar, biologically active GA is implied.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - GA gibberellin(s) or gibberellin-like substances - GC-RC gas chromatography-radiochromatogram counting - HPLC high-presare liquid chromatography - Rt retention time - TLC thinlaver chromatography     

Lactose carrier protein of Escherichia coli. Structure and expression of plasmids carrying the Y gene of the lac operon

The previously described hybrid plasmid pC7 which carries lacI+O+delta(Z)Y+A+ on a 12.3 X 10(6)-Mr DNA fragment [Teather et al. (1978) Mol. Gen. Genet. 159, 239-248] was partially digested with the restriction endonuclease EcoRI under conditions reducing the recognition sequence to d(A-A-T-T) and ligated to the vector pB322. lac Y-carrying inserts of various sized (Mr 1.5-4.7 X 10(6)) were obtained. Hybrid plasmid pTE18 (2300-base-pair insert) carries part of the I (repressor) gene, the promotor-operator region, part of the Z (beta-galactosidase) gene, the Y (lactose carrier) gene and part of the A (transacetylase) gene. Upon induction of pTE18-harbouring strains the Y-gene product is expressed at a nearly constant rate for several generations and accumulates to a level of 12-16% of the total cytoplasmic membrane protein. Integration into the membrane leads to active carrier as judged by binding and transport measurements.     

Ultrastructure of cell wall and thylakoid membranes of the thermophilic cyanobacterium Synechococcus lividus under the influence of temperature shifts

The ultrastructure of the cell wall and the thylakoid membranes of the thermophilic cyanobacterium Synechococcus lividus was studied by freezefracture electron microscopy after temperature shifts. Different fracture faces of the outer, the cytoplasmic and the thylakoid membranes were demonstrated when the preparation-temperature was in the range of the optimal growth temperature at 52°C or after fixation at 52°C. In the outer membrane of the cell wall two fracture faces with holes and 7.5 nm intramembrane particles were detected. On both the outer (EF) and inner (PF) leaflet of the cytoplasmic membrane randomly distributed particles were demonstrated. The particle density on the PF-face was approx. three times that of the EF-face. The EF-face of the thylakoid membrane exposed rows of particles with an average diameter of 10 nm. The spacing between the particle rows was 35–50 nm. This regular particle arrangement on the EF-face was demonstrated only in a few cases. Mostly the intramembrane particles were distributed randomly on the thylakoid fracture faces. The particle density of thylakoids with a random distribution was approx. in the same range both on the EF-and PF-face. The EF-particles fall into four groups of 9,10,11, and 12.5 nm. The main particle class was the 10 nm class. The PF-face exposed smaller particles with two maxima at 8.5–9 nm and 10 nm. When Synechococcus lividus OH-53s was chilled to temperatures below 30–35°C before the freeze-etch preparation a phase transition took place after the temperature shift. On the fracture faces of the thylakoid and cytoplasmic membranes particle depleted areas occurred. The size of the areas were different in both membranes and dependent on the velocity of cooling. Contrary to Synechococcus lividus OH-53s in the mesophilic Synechococcus strain 6910 the phase transition point was 15°C. The lower phase transition point may be due to a higher content of unsaturated fatty acids.Dedicated to Prof. D. Peters (Hamburg) on the occasion of the 65th anniversary of his birthday     

Feinstruktur der Tarsal-Drüse vonSiro duricovius (Joseph) (Opiliones,Sironidae)

Zusammenfassung Alle Arten der Sironidae tragen als sekundäres männliches Geschlechtsmerkmal ein epidermales Drüsenorgan im letzten Tarsalglied des 4. Laufbeines. Das Sekret tritt dort dorsal, exponiert auf einer kanülenartigen Apophyse, dem Adenostyl, nach außen.BeiSiro duricorius (Joseph, 1868) besteht das komplexe Drüsenorgan aus zahlreichen funktionellen Einheiten, deren Sekret einem gemeinsamen Kanal zugeführt wird; er öffnet sich auf dem Adenostyl. Jede funktionelle Drüseneinheit umfaßt 5 Zellen: 3 Drüsenzellen (DZ, Dreiergruppe), 1 Hüllzelle (HZ), 1 Kanalzelle (KZ). Die Sekrete werden in ein gemeinsames Reservoir abgegeben.HZ verknüpft manschettenartig die DZ mit der KZ. Eine Randeinfaltung am Apex der DZ gewährleistet innige Verfalzung von diesen mit der HZ.DZ und HZ sind mit gut entwickeltem Mikrovillisaum ausgestattet; beide Zelltypen sezernieren.Der ableitende Kanal der GZ besteht aus 2 Abschnitten: ein distaler schwammig-fibrillärer und ein proximaler massiver; beide Teile werden von nur einer Zelle (KZ) begleitet. Der basale Teil mündet in den Hauptkanal, der zum Adenostyl zieht.Neben diesem komplexen Drüsenorgan liegt ein ähnlich gebautes, aber kleineres ventral im Tarsus; sein Sammelkanal zieht zum Kanal der Hauptdrüse.Isolierte funktionelle Einheiten sind in der Tarsenepidermis weit verstreut; sie leiten ihr Sekret unmittelbar auf die Tarsusoberfläche.Die DZ produzieren nach elektronenoptischer Evidenz ein Protein. Das Sekret der HZ läßt sich bisher nicht zuordnen; es ist von dem der DZ verschieden.

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Ultrastructure of the tarsal gland ofSiro duricorius (Joseph)

Summary All species of Sironidae possess as a secondary male sex character an epidermal gland organ in the last tarsal joint of the fourth leg. The glandular secretion emanates from a conical apophysis, the adenostyl, which is located dorsally on the joint.InSiro duricorius (Joseph, 1868) the complex organ consists of numerous functional units, the secretion of which flows into the common channel opening on the adenostyl.Every gland unit contains 5 cells: 3 gland cells (DZ), 1 enveloping cell (HZ), 1 duct cell (KZ). The enveloping cell connects the gland cells with the duct cell like a sleeve. A marginal fold of the gland cell apex provides tight connection of gland cells and enveloping cell.Gland cells and enveloping cell are equipped with microvilli, and both cell types secrete in a common reservoir.The duct of the duct cell is divided into two parts: 1) a distal fibrillaceous-spongy one and 2) a proximal massive one. Both are accompanied only by a single duct cell.Besides this complex gland organ a similar one is located in the ventral part of the tarsus. Its collecting channel opens into the channel of the main gland.Isolated functional gland units are scattered over the epidermis of the tarsus; their secretion flows immediately on the tarsal surface.The gland cells proper produce a protein, the fact is indicated by the electronmicrographs. The secretion of the enveloping cell is a different one, but at this time cannot be attached to a chemical group.

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Abkürzungen A Adenostyl - aSe austretendes Sekret - C Kutikulin - Cu Kutikula - dK distaler Kanal - DK Kern der Drüsenzelle - DZ Drüsenzelle - ER rauhes endoplasmatischesm, Retikulum - G Golgi-Apparat - HK Kern der Hüllzelle - HZ Hüllzelle - K Kralle des Tarsus - Ke Kern - KK Kern der Kanalzelle - KZ Kanalzelle - M Mikrovilli - Mi Mitochondrium - pK proximaler Kanal - R Randeinfaltung der DZ - Re extrazelluläres Reservoir - Ri Ribosomen - Se Sekret - SK zentraler Sammelkanal - T Mikrotubuli - Va Vakuole - Va₁-Va₆ Vakuolen mit Sekret unterschiedlichen Reifegrades     

Formation of mesodermal pattern by secondary inducing interactions

Summary A highly purified vegetalizing factor induces endoderm preferentially in amphibian gastrula ectoderm. After combination of this factor with less pure fractions, a high percentage of trunks and tails with notochord and somites are induced. The induction of these mesodermal tissues depends on secondary factors which may act on plasma membrane receptors of the target cells. The secondary factors are probably proteins as they are inactivated by trypsin or cellulose-bound proteinase K. They are not inactivated by thioglycolic acid.The implication of these findings for tissue determination and differentiation in normal development in relation to the anlageplan for endoderm and mesodermal tissues is discussed.     

Genes inhibiting senescence in the ascomycete Podospora anserina.

Summary Senescence occurs in all wild strains of Podospora anserina after continued growth. This syndrome can be inhibited by a synergistic interaction of two linked genes, incoloris and vivax. Whereas the wild strain starts to become senescent after 26 d and the mutants incoloris and vivax after 42 and 66 d respectively, the double mutant shows no signs of aging after culture for more than one year.     

Griseofulvin interacts with microtubules both in vivo and in vitro

Immunofluorescence microscopy using monospecific tubulin antibody shows that in vivo griseofulvin interferes with the expression of both cytoplasmic and spindle microtubules in tissue culture cells in a concentration-dependent manner. In mouse 3T3 cells cytoplasmic microtubules are destroyed at a griseofulvin concentration of 5 × 10^?5m. At this concentration no increase of the mitotic index is observed but the cells are arrested in interphase, probably due to the destruction of cytoplasmic microtubules. Lowering the drug concentration to 10^?5m allows 3T3 cells to accumulate in c-mitotic (“colchicin-mitotic”) arrest. In HeLa cells the display of spindle microtubules observed in drug-arrested cells appears similar to that seen in normal metaphase cells only at lower griseofulvin concentrations. Higher drug concentrations induce c-mitotic arrest accompanied by an increasing loss of typical metaphase tubulin structures.In vitro polymerization experiments with brain tubulin using both light-scattering and electron microscopy show that in the presence of griseofulvin tubulin can aggregate rapidly in the cold. This behaviour is not found in the absence of the drug. Thus both in vivo and in vitro experiments show that griseofulvin, like other c-mitotic drugs, acts at the level of tubulin polymerization and that its effects are concentration dependent.     

Akustische Differenzierung verwandtschaftlicher Beziehungen in derParus (Periparus)-Gruppe nach Untersuchungen im Nepal-Himalaya

Zusammenfassung Die fünf Formen der UntergattungParus (Periparus) des südlichen Zentral-Asien(rufonuchalis, rubidiventris, beavani, melanolophus, aemodius) werden morphologisch, biologisch-ökologisch und vor allem akustisch differenziert. Die Spezifität der Gesänge gilt als wesentliches Art-Kriterium; sie wird mit Attrappen-Versuchen getestet.1.Rufonuchalis, rubidiventris undbeavani (Fichtenmeisen-Gruppe) bilden im Nepal-Himalaya Arealgrenzen aus.Rufonuchalis undrubidiventris leben dort sympatrisch,rubidiventris undbeavani allopatrisch ohne Introgressionszone. Rubidiventris undbeavani haben übereinstimmende ökologische Ansprüche;rufonuchalis als größere und trockenadaptierte Form weicht merklich ab. Die Arealgrenzen vonrufonuchalis undrubidiventris sind im westlichen Nepal-Himalaya klimatisch bedingt (Monsun!), nicht durch Konkurrenz mit anderenPeriparus-Formen.Reviergesänge der farblich fast ununterscheidbarenrufonuchalis undbeavani divergieren sehr, die vonrubidiventris undbeavani sind gleich. Rufonuchalis verfügt über 2 Gesangstypen: Trillergesang dient der Reviermarkierung, Pfiffgesang der Revierverteidigung. Beide Gesangsformen entsprechen nicht demParus-Gesangsschema, wohl aber der Klappergesang vonrubidiventris undbeavani. Beavani reagiert im Attrappen-Test nur auf denrufonuchalis-Pfiffgesang und besitzt vergleichbare Lautäußerungen.Rufonuchalis-Trillergesang findet beibeavani ebenfalls Entsprechungen, jedoch nicht beirubidiventris.Spezialisierte Trillergesang-Elemente vonrufonuchalis undbeavani werden als homolog betrachtet. Bei diesen allopatrischen Formen konnten sie erhalten bleiben, der mitrufonuchalis sympatrischerubidiventris hat sie reduziert.Die gleichfarbigenrufonuchalis undbeavani haben das Himalaya-System nie kontinuierlich besiedelt; ihre Areale standen ursprünglich über die Waldzonen Zentralasiens miteinander in Verbindung.Akustische Analyse und Verbreitungsmuster weisenrufonuchalis undrubidiventris (mit den Subspeziesrubidiventris undbeavani) als eigene Spezies aus; sie werden zu einer Superspezies verklammert.2. Die Areale vonP. ater aemodius undP. melanolophus stehen im zentralen Himalaya in sekundärem Kontakt; sie bilden dort eine Hybrid-(Introgressions-)Zone von 70 km Breite.An keiner Stelle desaemodius- bzw.melanolophus-Areals kommen die beiden Ausgangsformen bzw. Hybriden und Ausgangsformen nebeneinander vor. Merkliche Zuwanderung in das Hybridgebiet aus östlichen oder westlichen Arealteilen findet nicht statt.Es existieren Hybriden, die nicht intermediär zwischen den Ausgangsformen stehen, sondern von ihnen in Farbmuster und Färbung markant abweichen (rostbäuchige Hybriden).Vom Gebiet reineraemodius-Phäne zu dem reinermelanolophus-Phäne verläuft eine Merkmalsprogression:aemodius-Beige der Unterseite zu Rostrot;aemodius-Beige der Flanken zumelanolophus-Grau. Rostrot der Unterseite schlägt fast ohne Übergang zumelanolophus-Grau um (bisher nur für eine Population belegt).Alle Populationen des Hybridgürtels sind voll fortpflanzungsfähig (Reviergesang, Revierabgrenzung, -Brutfleck, -Gonadenentwicklung). Die Ausgangsformen haben (noch) keine verschiedenen adaptiven Niveaus erreicht; die Hybriden sind bei der Paarbildung (offensichtlich) nicht unterlegen.Isolationsmechanismen zwischen Ausgangsformen und Hybriden sind (heute) nicht (mehr) entwickelt; füraemodius undmelanolophus werden gering entwickelte akustische (und optische) postuliert (Hinweise durch Hybridisation in Gefangenschaft).Rostfarbigkeit wird als altes Merkmal derPeriparus-Gruppe angesehen, sein Auftreten bei den Hybriden als Atavismus.Reviergesänge aus derater/melanolophus-Gruppe sind einander sehr ähnlich. Der Gesang mitteleuropäischera. ater und der vonmelanolophus zeichnet sich durch ansteigende Elemente aus, diea. aemodius fehlen. Auf solche Gesangstypen reagiertaemodius nur mangelhaft.Die Gesänge in den Hybridpopulationen gleichen jenen vonaemodius; das wird auf Lernen desaemodius-Repertoires der Hybriden während des ersten Kontaktes vonmelanolophus undaemodius zurückgeführt.Süddeutschea. ater reagieren nur zur Zeit höchster Revierverteidigungs-Bereitschaft auf einen bestimmtenaemodius-Strophentyp. Auf diesen Strophentyp reagierten auch syntop lebendeP. major; er enthält Merkmale der Gesänge beider Arten.Die genetische Divergenz vonaemodius undmelanolophus — kenntlich an den rostbäuchigen Hybriden — legt nahe, den Artstatus beider bedingt aufrechtzuerhalten: SuperspeziesP. ater mit den SemispeziesP. ater undP. melanolophus.

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Acoustic differentiation of phylogenetic relations in theParus (Periparus) group from investigations in the Nepal Himalayas

Summary In the Nepal Himalayas 5 taxa of the subgenusParus (Periparus) exist — the greatest concentration of this subgenus in the whole Palearctic region. The taxa belong to 2 groups: 1.rufonuchalis, rubidiventris, beavani; 2. aemodius, melanolophus (fig. 1). All forms settle in the same altitude belt from about 2800 to 4200 m — three of them even in the same locality.Rufonuchalis is adapted to dry habitats mainly of West Central Asia and is thus restricted in Nepal to the Western parts (fig. 3, 4).The up to now unclarified status of all forms is determined as follows: Distinct species areP. rufonuchalis andP. rubidiventris (with highly different subspeciesP. r. rubidiventris andP. r. beavani) and they are regarded as a superspecies.P. rufonuchalis andP. r. rubidiventris live syntopically in West Nepal and display different territorial songs.Both differ remarkably in body size and bill size. Certain acoustic similarities indicating close relationship could be shown betweenrufonuchalis and the allopatric (fig. 3, 4)rubidiventris beavani; r. beavani reacts to the playback of whistle song (Pfiffgesang, fig. 10) ofP. rufonuchalis (see also fig. 14).The areas ofP. r. rubidiventris andP. r. beavani are separated apparently only by the Bhote Kosi river in East Nepal (fig. 4). Introgression is not (yet) known; the songs are identical (fig. 8). P. melanolophus andP. ater aemodius hybridize in West Nepal in a zone of secondary contact. Their areas are linked by a zone of introgression (fig. 17, 18). In that zone cinnamomeous bellied hybrids locally occur differing thus strikingly in colour from both parental species (fig. 1). Sympatric occurrence ofmelanolophus andater aemodius is not known and is not expected either.Territorial songs ofa. aemodius (fig. 21) andmelanolophus (fig. 25) are very similar and both species obviously do not differ in their ecology. Despite the small introgression belt, similar songs and apparently very similar or even identical adaptive peaks,P. ater andP. melanolophus are still treated as distinct species but reduced to semispecies level. In doing so the genetic divergence betweenater aemodius andmelanolophus indicated by the cinnamomeous bellied hybrids is emphasized.

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Mit einem Jahresstipendium des Deutschen Akademischen Austauschdienstes und einer Sachbeihilfe der Deutschen Forschungsgemeinschaft. — Ergebnisse der Nepal-Reisen 1969/70, 1973 und 1974, Nr. 36. — Nr. 35: Senckenbergiana biol., 56 (4/6): 247–256, 1975.