Molecular mechanisms of hormone controlled gene expression in the breast

In a meritorious effort H. de Rothschild compiled in 1899 all publications on mammary gland development and milk – a grand total of 8375 [1]. In the preface to this publication Duclaux states: ‘Such a discrepancy between the tremendous efforts and the paltriness of the results – hundreds of scientists and thousands of research years, just to create 200 or 300 pages of truth’. The number of papers added since then must be enormous. Rather than reviewing a vast literature, I will take the liberty and focus on research which, in my opinion, shaped our understanding of hormone controlled gene expression in the developing breast. This revised version was published online in August 2006 with corrections to the Cover Date.     

Jasmonic acid spraying does not induce tuberisation in short-day-requiring potato species kept in non-inducing conditions

The potato species Solanum andigena (Juz. and Buk.) and Solanum demissum (Lindl.) that both require short days for tuberisation were kept in either long days (16 h light), or short days (8 h light) with a 30-min night break mid-way through the dark period. Tuberisation of these species was inhibited under both conditions. Repeated spraying of these plants with up to 100 μM jasmonic acid did not induce them to tuberise even though jasmonic acid was shown to be taken up and transported within the plant. This result argues against jasmonic acid itself being the transported tuber-inducing signal, although it does not exclude a role for jasmonic acid later in tuber formation and development once induction has taken place.     

Isolation and characterization of two cDNA clones encoding ATP-sulfurylases from potato by complementation of a yeast mutant

Sulfur plays an important role in plants, being used for the biosynthesis of amino acids, sulfolipids and secondary metabolites. After uptake sulfate is activated and subsequently reduced to sulfide or serves as donor for sulfurylation reactions. The first step in the activation of sulfate in all cases studied so far is catalyzed by the enzyme ATP-sulfurylase (E.C. 2.7.7.4.) which catalyzes the formation of adenosine-5′-phosphosulfate (APS). Two cDNA clones from potato encoding ATP-sulfurylases were identified following transformation of a Saccharomyces cerevisiae mutant deficient in ATP-sulfurylase activity with a cDNA library from potato source leaf poly(A)⁺ RNA cloned in a yeast expression vector. Several transformants were able to grow on a medium with sulfate as the only sulfur source, this ability being strictly linked to the presence of two classes of cDNAs. The clones StMet3-1 and StMet3-2 were further analyzed. DNA analysis revealed an open reading frame encoding a protein with a molecular mass of 48 kDa in the case of StMet3-1 and 52 kDa for StMet3-2. The deduced polypeptides are 88% identical at the amino acid level. The clone StMet3-2 has a 48 amino acid N-terminal extension which shows common features of a chloroplast transit peptide. Sequence comparison of the ATP-sulfurylase Met3 from Saccharomyces cerevisiae with the cDNA StMet3-1 (StMet3-2) reveals 31% (30%) identity at the amino acid level. Protein extracts from the yeast mutant transformed with the clone StMet3-1 displayed ATP-sulfurylase activity. RNA blot analysis demonstrated the expression of both genes in potato leaves, root and stem, but not in tubers. To the best of the authors' knowledge this is the first cloning and identification of genes involved in the reductive sulfate assimilation pathway from higher plants.     

Riesenkerne im Endosperm vonAllium ursinum

Zusammenfassung In der chalazalen Region des Endosperms vonAllium ursinum entstehen Riesenkerne, die eine besondere permanent-prophasische Struktur besitzen und während ihres Wachstums polyploid, und zwar zunächst offenbar hexaploid und 12-ploid werden. Sie entwickeln sich völlig kontinuierlich, also ohne zwischengeschaltete Mitosen oder Interphasen, aus triploiden Kernen des jungen Endosperms, die selbst stark vergrößert sind. In alten Embryosäcken treten auch Riesenkerne auf, die mit größter Wahrscheinlichkeit als 24-ploid anzusprechen sind.Die Polyploidisierung der Riesenkerne entspricht nicht dem für Angiospermen typischen endomitotischen Formwechsel. Sie ist wohl als besonders extreme Hemmung und Umbildung von Mitosen aufzufassen, die eine Eigentümlichkeit des Endosperms darstellt.Die 12-ploiden Riesenkerne besitzen gegenüber diploiden Kernen des äußeren Integuments ungefähr das 100fache Volumen, gegenüber den vermutlich oktoploiden Kernen des Wassergewebes der Samenanlagen ungefähr das 20–30fache, gegenüber den triploiden Kernen des jungen Endosperms das 4–7fache Volumen. Die bedeutende Größe der triploiden Kerne des jungen Endosperms —später werden die Kerne sukzessive kleiner —beruht auf gesteigerter Chromosomengröße (die Chromosomen sind etwa 3 1/2 mal voluminöser als in anderen Mitosen) und auf Vermehrung extrachromosomaler Substanz.Die Kerne des Wassergewebes und des gewöhnlichen Endosperms besitzen gewebespezifische Struktur; die Riesenkerne des physiologisch hoch aktiven Basalapparates sind besonders auffallend gewebespezifisch organisiert: sie enthalten in prophasischer Ausbildung Diplo-, bzw. Quadruplound vermutlich Oktuplochromosomen.Als gelegentliche Beobachtungen an nichtchalazalen Regionen des Endosperms seien erwähnt: das Auftreten von Spindeln, die an einem Pol einen, am anderen Pol zwei Tochterzellen ergeben; das Vorkommen häufiger Mitosestörungen in späteren Stadien und wohl damit im Zusammenhang die Bildung hexaploider und wohl 12-ploider Endosperm-kerne, die weitere Mitosen einzugehen vermögen.     

Lebendbeobachtung der Chromatophorenteilung der DiatomeeNitzschia

Continuous intra vitam observations inNitzschia palea and supplementary ones in two otherN.-species show that the division of the platelike chromatophore is intimately connected with mitosis and cytokinesis. It starts and goes on in a strictly regular course. When the ingrowing plasmatic zone of cytokinetic separation reaches the distal edge of each chromatophore, a narrow split originates and deepens until it has reached the middle of the chromatophore or a little more. Only then a second split arises at the proximal edge of the chromatophore; from there it grows in inverse direction, until it meets the first formed split in a distance of about one third from the proximal end of the chromatophore. Under normal circumstances the position of the meeting point is nearly invariable, the speed of the two progressing splits is the same, and the splitting of the two chromatophores of one cell is nearly exactly synchronized. There is only little variation in the beginning, as the split can form in somewhat different ways. Thereby a striking plasticity of the chromatophore is apparent. The cytokinesis including chromatophore division lasts about 4 1/2–5 minutes. Until now the mode of chromatophore division inNitzschia remains without analogy.     

Das Sediment des Kummerower Sees. Untersuchungen des Chemismus und der Diatomeenflora

The Sediment of Lake Kummerow. Investigations on the Chemism and the Diatom Flora The paper deals with the distribution of chemical parameters and of diatoms in the deposits of the eutrophic Lake Kummerow (GDR), produced during about the last 3500 years. The sediment consists of a calcareous gyttja with a very low content of sand and clay. The results indicate a relatively constant trophic state of the lake in former times and an increased production within the last five centuries. Stephanodiscus astraea and its variety minutula is by far the most constant diatom species; in the uppermost parts of the sediment small sized planctonic taxa are increasing, in the main Stephanodiscus hantzschii var. pusillus, Melosira granulata var. angustissima, Asterionella formosa and Synedra acus var. angustissima and var. radians.     

High-Resolution Metabolic Phenotyping of Genetically and Environmentally Diverse Potato Tuber Systems. Identification of Phenocopies

We conducted a comprehensive metabolic phenotyping of potato (Solanum tuberosum L. cv Desiree) tuber tissue that had been modified either by transgenesis or exposure to different environmental conditions using a recently developed gas chromatography-mass spectrometry profiling protocol. Applying this technique, we were able to identify and quantify the major constituent metabolites of the potato tuber within a single chromatographic run. The plant systems that we selected to profile were tuber discs incubated in varying concentrations of fructose, sucrose, and mannitol and transgenic plants impaired in their starch biosynthesis. The resultant profiles were then compared, first at the level of individual metabolites and then using the statistical tools hierarchical cluster analysis and principal component analysis. These tools allowed us to assign clusters to the individual plant systems and to determine relative distances between these clusters; furthermore, analyzing the loadings of these analyses enabled identification of the most important metabolites in the definition of these clusters. The metabolic profiles of the sugar-fed discs were dramatically different from the wild-type steady-state values. When these profiles were compared with one another and also with those we assessed in previous studies, however, we were able to evaluate potential phenocopies. These comparisons highlight the importance of such an approach in the functional and qualitative assessment of diverse systems to gain insights into important mediators of metabolism.     

Structure of a highly phosphorylated lipopolysaccharide core in the Delta algC mutants derived from Pseudomonas aeruginosa wild-type strains PAO1 (serogroup O5) and PAC1R (serogroup O3)

Lipopolysaccharides (LPS) were isolated from rough-type mutant strains of Pseudomonas aeruginosa (Delta algC) derived from wild-type strains PAO1 (serogroup O5) and PAC1R (serogroup O3). Structural studies of the LPS core region with a special focus on the phosphorylation pattern were performed by 2D NMR spectroscopy, including a 1H,(31)P HMQC-TOCSY experiment, MALDI-TOF MS, and Fourier-transform ion cyclotron resonance ESIMS using the capillary skimmer dissociation technique. Both LPS were found to contain two residues each of 3-deoxy-D-manno-oct-2-ulosonic acid (Kdo) and L-glycero-D-manno-heptose (Hep), one residue of N-(L-alanyl)-D-galactosamine and one O-carbamoyl group (Cm) on the distal Hep residue. The following structures of a tetrasaccharide trisphosphate from strain PAC1R Delta algC and that carrying an additional ethanolamine phosphate group (PEtN) from strain PAO1 Delta algC were elucidated: [carbohydrate structre: see text] where R=P in PAC1R Delta algC and PPEtN in PAO1 Delta algC. To our knowledge, in this work the presence of ethanolamine diphosphate is unambiguously confirmed and its position established for the first time in the LPS core of a rough-type strain of P. aeruginosa. In addition, the structure of the complete LPS core of wild-type strain P. aeruginosa PAO1 was reinvestigated and the position of the phosphorylation sites was revised.     

Structure-function relationship studies of PTH(1-11) analogues containing sterically hindered dipeptide mimetics.

The N-terminal 1-34 fragment of parathyroid hormone (PTH) is fully active in vitro and in vivo and reproduces all biological responses characteristic of the native intact PTH. In order to develop safer and non-parenteral PTH-like bone anabolic agents, we have studied the effect of introducing conformationally constrained dipeptide mimetics into the N-terminal portion of PTH in an effort to generate miniaturized PTH-mimetics. To this end, we have synthesized and conformationally and biologically characterized PTH(1-11) analogues containing 3R-carboxy-6S-amino-7,5-bicyclic thiazolidinlactam (7,5-bTL), a rigidified dipeptide mimetic unit. The wild type sequence of PTH(1-11) is H-Ser-Val-Ser-Glu-Ile-Gln-Leu-Met-His-Asn-Leu-NH(2). The following pseudo-undecapeptides were prepared: [Ala(1), 7,5-bTL(3, 4), Nle(8), Arg(11)]hPTH(1-11)NH(2) (I); [Ala(1), 7,5-bTL(6, 7), Nle(8), Arg(11)]hPTH(1-11)NH(2) (II); [Ala(1), Nle(8), 7,5-bTL(9, 10), Arg(11)]hPTH(1-11)NH(2) (III). In aqueous solution containing 20% TFE, only analogue I exhibited the typical CD pattern of the alpha-helical conformation. NMR experiments and molecular dynamics calculations located the alpha-helical stretch in the sequence Ile(5)-His(9). The dipeptide mimetic unit 7,5-bTL induces a type III beta-turn, occupying the positions i - 1 and i of the turn. Analogue II exhibited an equilibrium between a type I beta-turn and an alpha-helix, and analogue III did not show any ordered structure. Biological tests revealed poor activity for all analogues (EC(50) > 0.1 mM). Apparently, the relative side-chain orientation of Val(2), Ile(5) and Met(8) can be critical for effective analogue-receptor interaction. Considering helicity as an essential property to obtain active PTH agonists, one must decorate the correctly positioned dipeptide mimetic azabicycloalkane scaffold with substitutions corresponding to the displaced amino acids.