Cloning of aroG, the gene coding for phospho-2-keto-3-deoxy-heptonate aldolase(phe), in Escherichia coli K-12, and subcloning of the aroG promoter and operator in a promoter-detecting plasmid

Defective transducing phages carrying aroG, the structural gene for phenylalanine (phe)-inhibitable phospho-2-keto-heptonate aldolase (EC 4.1.2.15; previously known as 3-deoxy-D-arabinoheptulosonate-7-phosphate synthetase[phe]), have been isolated, and DNA from two of these phages has been used to construct a restriction map of the region from att lambda to aroG. A 7.6-kb PstI-HindIII fragment from one of these phages was cloned into pBR322 and shown to contain aroG. The location of aroG within the 7.6 kb was established by subcloning and Tn3 transpositional mutagenesis. A fragment carrying the aroG promoter and operator has been cloned into a high copy number promoter-cloning vector (pMC489), and the resulting aroGpo-LacZ' (alpha) fusion subcloned in a low copy number vector. Strains with this fusion on the low copy number vector exhibit negative regulation of beta-galactosidase expression by both phenylalanine and tryptophan and positive regulation by tyrosine in a tyrR+ background.     

Evaluation of four-dimensional cone beam computed tomography ventilation images acquired with two different linear accelerators at various gantry speeds using a deformable lung phantom

We evaluated four-dimensional cone beam computed tomography (4D-CBCT) ventilation images (VI_CBCT) acquired with two different linear accelerator systems at various gantry speeds using a deformable lung phantom.The 4D-CT and 4D-CBCT scans were performed using a computed tomography (CT) scanner, an X-ray volume imaging system (Elekta XVI) mounted in Versa HD, and an On-Board Imager (OBI) system mounted in TrueBeam. Intensity-based deformable image registration (DIR) was performed between peak-exhale and peak-inhale images. VI_CBCT- and 4D-CT-based ventilation images (VI_CT) were derived by DIR using two metrics: one based on the Jacobian determinant and one on changes in the Hounsfield unit (HU). Three different DIR regularization values (λ) were used for VI_CBCT. Correlations between the VI_CBCT and VI_CT values were evaluated using voxel-wise Spearman’s rank correlation coefficient (r).In case of both metrics, the Jacobian-based VI_CBCT with a gantry speed of 0.6 deg/sec in Versa HD showed the highest correlation for all the gantry speeds (e.g., λ = 0.05 and r = 0.68). Thus, the r value of the Jacobian-based VI_CBCT was greater or equal to that of the HU-based VI_CBCT. In addition, the ventilation accuracy of VI_CBCT increased at low gantry speeds.Thus, the image quality of VI_CBCT was affected by the change in gantry speed in both the imaging systems. Additionally, DIR regularization considerably influenced VI_CBCT in both the imaging systems. Our results have the potential to assist in designing CBCT protocols, incorporating VI_CBCT imaging into the functional avoidance planning process.     

Functional lung imaging with synchrotron radiation: Methods and preclinical applications

Many lung disease processes are characterized by structural and functional heterogeneity that is not directly appreciable with traditional physiological measurements. Experimental methods and lung function modeling to study regional lung function are crucial for better understanding of disease mechanisms and for targeting treatment. Synchrotron radiation offers useful properties to this end: coherence, utilized in phase-contrast imaging, and high flux and a wide energy spectrum which allow the selection of very narrow energy bands of radiation, thus allowing imaging at very specific energies. K-edge subtraction imaging (KES) has thus been developed at synchrotrons for both human and small animal imaging. The unique properties of synchrotron radiation extend X-ray computed tomography (CT) capabilities to quantitatively assess lung morphology, and also to map regional lung ventilation, perfusion, inflammation and biomechanical properties, with microscopic spatial resolution. Four-dimensional imaging, allows the investigation of the dynamics of regional lung functional parameters simultaneously with structural deformation of the lung as a function of time. This review summarizes synchrotron radiation imaging methods and overviews examples of its application in the study of disease mechanisms in preclinical animal models, as well as the potential for clinical translation both through the knowledge gained using these techniques and transfer of imaging technology to laboratory X-ray sources.     

Activation of fatty acids by non-glyoxysomal peroxisomes

Ethylene treatment (approx. 20 l ·1^-1 in air for 2 d) of tobacco (Nicotiana tabacum L. cv. Havana 425) plants markedly increases the endo--1,3-glucanase (EC 3.2.1.39) content of leaves. The antigenic form of the enzyme induced is the same one whose production is blocked by treating cultured cells with combinations of auxin (1.1 · 10^-5 M -naphthaleneacetic acid) and cytokinin (1.4 · 10^-6 M kinetin). Evidence is presented that cultured tobacco cells require ethylene for -1,3-glucanase accumulation: i) ethylene treatment increased the accumulation of \-1,3-glucanase in callus tissues >10 d after subculturing and in cell-suspension cultures; ii) callus tissues can produce ethylene; iii) conditions known to inhibit ethylene production (1 mM CoCl₂; 33° C treatment) or ethylene action (approx. 1.6 mmol · 1^-1 norbornadiene in air) inhibited -1,3-glucanase accumulation by callus tissues treated for 4 d following subculturing; and, these inhibitory effects were prevented by exogenous ethylene. Combinations of auxin and cytokinin blocked ethylene-induced accumulation of -1,3-glucanase by cell-suspension cultures. The results favor a model in which ethylene induces results favor a model in which ethylene induces 1,3-glucanase accumulation, and auxin and cytokinin inhibit this induction process.Abbreviations NAA -naphthaleneacetic acid - NDE norbornadiene     

C-banding polymorphism and the analysis of nucleolar activity in Dasypyrum villosum (L.) Candargy,its added chromosomes to hexaploid wheat and the amphiploid Triticum dicoccum — D. villosum

Summary C-banding patterns and nucleolar activity were analyzed in Dasypyrum villosum, its added chromosomes to hexaploid wheat and the hexaploid amphiploid Triticum dicoccum-D. villosum. Two different populations of the allogamous species D. villosum (2n= 14, VV) from Greece and Italy were analyzed showing a similar polymorphism for C-banding pattern. Six of the seven addition lines were identified by their characteristic C-banding pattern. No polymorphism between both members of each added alien chromosome was found. Furthermore, nucleolar activity and competition were studied by using silver staining procedure. In D. villosum only one chromosome pair, A, was found to be responsible for organizing nucleoli. The results obtained in the amphiploid and in the addition lines demonstrate that nucleolar activity is restricted to SAT-chromosomes 1B and 6B of wheat, while those of D. villosum remain inactive.     

Organization of Methanobacterium thermoautotrophicum bacteriophage psi M1 DNA

Summary M1 is a virulent bacteriophage of Methanobacterium thermoautotrophicum strain Marburg. Restriction enzyme analysis of the linear, 30.4 kb phage DNA led to a circular map of the 27.1 kb M1 genome. M1 is thus circularly permuted and exhibits terminal redundancy of approximately 3 kb. Packaging of M1 DNA from a concatemeric precursor initiates at the pac site which was identified at coordinate 4.6 kb on the circular genome map. It proceeds clockwise for at least five packaging rounds. Headful packaging was also shown for M2, a phage variant with a 0.7 kb deletion at coordinate 23.25 on the map.     

Synthesis and conformational analysis of aib-containing peptide modelling for N-glycosylation site in N-glycoprotein

A tetrapetide containing an Aib residue, Boc-Asn-Aib-Thr-Aib-OMe, was synthesized as a peptide model for the N-glycosylation site in N-glycoproteins. Backbone conformation of the peptide and possible intramolecular interaction between the Asn and Thr side chains were elucidated by means of n.m.r. spectroscopy. Temperature dependence of NH proton chemical shift and NOE experiments showed that Boc-Asn-Aib-Thr-Aib-OMe has a tendency to form a β-turn structure with a hydrogen bond involving Thr and Aib⁴ NH groups. Incorporation of Aib residues in the peptide model promotes folding of the peptide backbone. With folded backbone conformation, carboxyamide protons of the Asn residue are not involved in hydrogen bond network, while the OH group of the Thr residue is a candidate for a hydrogen bond in DMSO-d₆ solution.     

Cellular origin and distribution of transforming growth factor-β1 in the normal rat myocardium

Summary Transforming growth factor- (TGF-) is a biologically active polypeptide present in normal tissues as well as transformed cells. Two structurally related forms of this peptide are TGF- ₁ and TGF- ₂. Using freshly isolated cardiomyocytes and non-myocyte heart cells, and a [³²P]-labelled cDNA probe to human TGF- ₁, we demonstrated that mRNA for TGF- ₁ could be detected only in the nonmyocyte fraction of heart cells. In the present study, the distribution of TGF- ₁ in the heart was determined by immunofluorescence staining by use of a polyclonal antibody to porcine TGF- ₁ in cryostat sections of rat heart. Immunofluorescence staining was intense around the blood vessels and radially diffuse in the surrounding myocardium.     

Characterization of the follicles in the intermediate lobe of the pituitary gland of the Mongolian gerbil (Meriones unguiculatus)

Summary The Mongolian gerbil (Meriones unguiculatus) contains abundant follicles throughout the intermediate lobe (IL) of the pituitary gland in the adult animal. The mode of follicle formation, the nature of the follicle building cells and the distribution of follicles were investigated in semithin sections of the gerbil IL. The sections were stained conventionally, or immunohistochemically with antibodies directed against -melanocyte stimulating hormone (- MSH). Follicular cells were constantly -MSH-negative, and resembled the marginal cells lining the hypophyseal cleft with regard to their cytological and immunohistochemical properties. Moreover, follicular cells appeared to be derived from strands of marginal cells that regularly invaginated deep into the IL. Both follicular and marginal cells often made up cellular clusters. This process coincided with follicle formation and the generation or transport of the colloidal content found inside follicles and the hypophyseal cleft. Although the non-secretory cells of the IL obviously constituted one major source of pituitary colloid in the gerbil, -MSH-positive secretory cells, which occasionally were found to be discharged into the cleft cavity, might contribute to the colloidal contents.     

Les régulateurs de croissance d'insectes (RCI), mimiques de l'hormone juvénile,en tant que moyen de lutte morphogénétique et ovicide contre les tordeuses des vergers

Résumé La métamorphose des insectes est régie par un équilibre hormonal complexe dans lequel l'hormone juvénile (HJ) joue un rôle important. Au dernier stade larvaire, la teneur en HJ est particulièrement faible dans le corps de l'insecte. Si un régulateur de croissance d'insectes (RCI)-un mimétique de l'HJ-est appliqué à ce moment-là, la mue nymphale est pertubée provoquant des déformations morphogénétiques caractéristiques. La teneur en HJ est également très faible dans les ufs fraîchement pondus. Les traitements aux RCI peuvent par conséquent perturber le développement embryonnaire de certaines espèces et produire ainsi un effet ovicide. Depuis quelques années deux RCI-le fenoxycarb et le CGA 45 128-ont été testés pour leur activité morphogénétique sur le dernier stade larvaire de quelques ravageurs tels qu'Adoxophyes orana F.v.R., ainsi que pour leur activité ovicide sur les ufs frais de Cydia pomonella L. et Grapholita funebrana Tr. Après quelques années d'expérimentation et de commercialisation des RCI dans les vergers européens, il s'avère que l'utilisation de ces produits peu toxiques, sélectifs et peu nocifs pour la faune utile, constitue une amélioration considérable pour l'aménagement de la lutte intégrée.

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Insect growth regulators (IGR), mimics of juvenile hormone, as morphological and ovicidal means of control against orchard tortricids

Summary Metamorphosis is regulated by a complex hormonal balance in which juvenile hormone (JH) plays an important part. At the last larval instar the content of JH is particularly low in the insect body. If an insect growth regulator (IGR) — a mimic of JH-is applied at this time, the pupal moult may be disturbed with the characteristic morphogenetical deformations. The JH content is also very low in freshly laid eggs. Therefore IGR treatments may disturb the embryonic development of some species and produce an ovicidal activity. During a few years two IGR-fenoxycarb and CGA 45128-were evaluated for their morphogenetical effect on the last larval instar of Adoxophyes orana F.v.R. and their ovicidal effect on freshly laid eggs of Cydia pomonella L. and Grapholita funebrana Tr. After a few years of experimentation with both compounds and of commercialisation of fenoxycarb in European orchards, IGR confirmed to present a considerable improvement in integrated pest management due to selectivity, and low mammal toxicity.