Evidence that replication initiates at only some of the potential origins in each oligomeric form of bovine papillomavirus type 1 DNA.

In a subclone of ID13 mouse fibroblasts latently infected with bovine papillomavirus type 1 (BPV-1) DNA, the viral genome occurred as a mixture of extrachromosomal circular monomers and oligomers. Multiple copies were also associated with the host cell genome, predominantly at a single site in a head-to-tail tandem array. We examined the replicative intermediates of extrachromosomal forms of BPV-1 DNA by using two-dimensional gel electrophoresis. The results obtained indicate that initiation of DNA replication occurred near the center of the EcoRI-BamHI 5.6-kilobase fragment. In some molecules, however, this fragment was replicated from one end to the other by means of a single fork initiated elsewhere. Termination also occurred within this fragment. The EcoRI-BamHI 2.3-kilobase fragment replicated as a DNA molecule containing a termination site for DNA replication and also by means of a single fork traversing the fragment from one end to the other. Thus, replication forks proceeded through these fragments in different manners, apparently depending on whether they were part of a monomer, a dimer, a trimer, or higher oligomers. These observations lead to the conclusion that initiation of DNA replication in BPV-1 DNA takes place at or close to plasmid maintenance sequence 1. From this point, replication proceeds bidirectionally and termination occurs approximately 180 degrees opposite the origin. The results obtained are consistent with one or more replication origins being quiescent in BPV-1 DNA oligomers.     

Effect of pectin on pectinases in autolysis of Botrytis cinerea

Pectic activity in autolyzed cultures of Botrytis cinerea in a medium with and without pectin was similar, but in the medium with pectin maximal activities occurred in younger cultures. The pectic activities found were polygalacturonase, polymethylgalacturonase, endo activity (pectin as substrate) and pectin lyase. The molecular weights of polygalacturonase, polymethylgalacturonase and endo activity (pectin as substrate) were 36000, 33000 and 30200 daltons respectively, and the molecular weight of pectin lyase was 18200 daltons. By gel electrophoresis four different pectic activities were detected, three in the top of the gel and one in the bottom. Two enzymes were characterized, the polygalacturonase activity (first band in the top) inhibited by Ca⁺⁺ and the pectin lyase activity (in the bottom) which was not inhibited by Ca⁺⁺. These enzymes are not induced by the presence of pectin in the medium during degradation of Botrytis cinerea.     

Isolation and biochemical characterization of Streptomyces clavuligerus mutants in the biosynthesis of clavulanic acid and cephamycin C

Summary Seven mutants of Streptomyces clavuligerus blocked in the biosynthesis of clavulanic acid, cephamycin C, or both antibiotics, have been isolated and characterized. Mutants nca1 and nca2 were unable to synthesize clavulanic acid but produced cephamycin C. Mutants nce1 and nce2 were completely blocked in cephamycin C production but formed clavulanic acid. A third group (mutants ncc1, ncc4 and ncc5) failed to produce both antibiotics. Arginase activity (forming ornithine) was very low in mutants ncc1 and ncc5. All the mutants blocked in clavulanic acid biosynthesis showed a normal ornithine--aminotransferase activity. Mutant ncc1, blocked in cephamycin biosynthesis, lacked completely lysine--aminotransferase (forming -aminoadipic acid) and isopenicillin N synthase. Two other mutants (nce2 and nce5) lacked isopenicillin N synthase. There was a good correlation between the isopenicillin N synthase and the lysine--aminotransferase activities of the nca mutants and the ability of those strains to produce cephamycin C. The condensing enzyme involved in the formation of the clavulanic acid nucleus appears to be different from the isopenicillin N synthase.Dedicated to Professor H.-J. Rehm on the occasion of his 60th birthday     

Effects of social isolation and crowding upon adrenocortical reactivity and behavior in the rat

The effects of social isolation and crowding on adrenocortical function and upon behavioral responsiveness to electric shock have been studied in male and female rats. All female experimental groups showed higher corticosterone levels and heavier adrenals than their male counterparts. The major effect of housing condition concerned the corticosterone response to stress, while basal hormone concentration was not modified. Socially housed rats showed a more intense adrenocortical response and also a greater behavioral reactivity to electric shock than the isolates.     

Control of Nitrogenase mRNA Levels by Products of Nitrate Assimilation in the Cyanobacterium Anabaena sp. Strain PCC 7120

Nitrate inhibited nitrogenase synthesis and heterocyst development in the cyanobacterium Anabaena sp. strain PCC 7120. Inhibition of dinitrogen fixation by nitrate did not take place, however, in nitrate reductase-deficient derivatives of this strain. Hybridization of total RNA isolated from cells grown on different nitrogen sources with an internal fragment of the nifD gene showed that regulation of nitrogenase activity by nitrate is exerted through a negative control of the nitrogenase mRNA levels.     

Cytology and morphology of the amphiploid Hordeum chilense (4x) × Aegilops squarrosa (4x)

Summary The intergeneric amphiploid Hordeum chilense × Aegilops squarrosa has been synthesized. The amphiploid plants have the expected chromosome number of 28. The average meiotic chromosome pairing was 12.48 bivalents + 3.04 univalents. The morphology of the amphiploid resembles that of the Aegilops parent. Nucleoli from both H. chilense and A. squarrosa are expressed in the amphiploid. Neither chromosome instability nor homoeologous pairing was found. The amphiploid is fertile and vigorous.     

The plasma membrane of Entamoeba histolytica: structure and dynamics.

The plasma membrane components of the parasitic protozoan Entamoeba histolytica, the causative agent of human invasive amebiasis, have been biochemically and immunologically characterized during the last decade. In addition, genes coding for certain surface proteins have been cloned. In spite of these advances, a unified characterization of plasma membrane antigenic components of the parasite is still required for badly needed advancements in the design of useful diagnostic, epidemiologic, and immunoprophylactic tools. Here we review current knowledge on this issue and address the problem of the considerable variation in the electrophoretic profiles of plasma membrane proteins obtained by different groups. In addition, the differences in the degree of recognition of reported membrane antigens with human immune sera, and the diverse interpretations concerning the possible functions of the surface molecules characterized are discussed. A comparative analysis of plasma membrane proteins of E histolytica trophozoites using three different isolation methods revealed that it is possible to select for specific membrane proteins, depending on the lysis conditions. In our view, the method of Calderón and Avila preserves more proteins than other methods tested. Using sera from recent cases of invasive amebiasis studied by several laboratories in various geographical areas, a basic antigenic pattern of 11 principal proteins with molecular weights of 220, 170, 150, 125, 97, 80, 60, 45, 20 and 9 kDA was established for the pathogenic E histolytica strain HM1:IMSS, used by most research groups.     

Immunocytochemical and ultrastructural characterization of endocrine cells in chicken proventriculus

Summary The endocrine cells of the chicken proventriculus were investigated immunocytochemically, using the peroxidase-antiperoxidase technique on paraffin and semithin sections for light microscopy, and immunogold staining in osmium-fixed material for electron microscopy. The fixation procedure also allowed a detailed ultrastructural investigation. Twenty-three antisera were tested and 7 immunoreactive cell-types were identified: D-cells containing somatostatin-like peptide; EG-cells immunoreactive to anti-glucagon, anti-GLP1 and antineurotensin; NT-cells labelled only with anti-neurotensin; BN-cells containing bombesin-like material; ENK-cells showing met-enkephalin immunoreactivity; EC-cells reactive to anti-serotonin; and APP-cells positive to anti-avian pancreatic polypeptide. In addition, enterochromaffin-like (ECL) cells, were also detected by electron microscopy. The presence of ENK-cells and the ultrastructure of these and NT-cells are described for the first time in chicken proventriculus, and glucagon, GLP1 and neurotensin are shown to be colocalized in the EG-cells.     

Wide distribution of a 36-megadalton plasmid among clinical and environmental Spanish isolates ofLegionella pneumophila serogroup 1

With the eventual goal of characterizingLegionella pneumophila serogroup 1 plasmids at the molecular level, we have analyzed the plasmid contents of 78 clinical and environmental Spanish isolates. After selection of a suitable alkaline lysis method, we detected plasmids with approximate molecular weights of 25, 36, 40, 61, 80, 85, 90, and 95 megadalton (MDal). Several factors (i.e., wide temporal and geographic distribution, high frequency in both clinical and environmental isolates, and apparent high copy number after subculturing) make the 36 MDal type IA plasmid an appropriate plasmid for further molecular studies.     

The R-banding pattern of the Chinese hamster Don cell line

Chinese hamster cells (Don line) were treated in vivo with 5-BrdU and 33258-Hoechst fluorochrome for obtaining the partial inhibition of condensation that causes the R-banding pattern. Untreated chromosomes were stained by a standard G-banding method. Statistical measurements show significant differences in the band numbers between the two treatments. The Don cell line in the authors' laboratory presents some karyotypical differences from Don cell lines studied by other authors.