Nucleotide sequence of the fixABC region of Azorhizobium caulinodans ORS571: similarity of the fixB product with eukaryotic flavoproteins, characterization of fixX, and identification of nifW.

Summary The nucleotide sequence of a 4.1 kb DNA fragment containing the fixABC region of Azorhizobium caulinodans was established. The three gene products were very similar to the corresponding polypeptides of Rhizobium meliloti. The C-terminal domains of both fixB products displayed a high degree of similarity with the -subunits of rat and human electron transfer flavoproteins, suggesting a role for the FixB protein in a redox reaction. Two open reading frames (ORF) were found downstream of fixC. The first ORF was identified as fixX on the basis of sequence homology with fixX from several Rhizobium and Bradyrhizobium strains. The second ORF potentially encoded a 69 amino acid product and was found to be homologous to a DNA region in the Rhodobacter capsulatus nif cluster I. Insertion mutagenesis of the A. caulinodans fixX gene conferred a Nif^– phenotype to bacteria grown in the free-living state and a Fix^– phenotype in symbiotic association with the host plant Sesbania rostrata. A crude extract from the fixX mutant had no nitrogenase activity. Furthermore, data presented in this paper also indicate that the previously identified nifO gene located upstream of fixA was probably a homologue of the nifW gene of Klebsiella pneumoniae and Azotobacter vinelandii.     

Polyester Wadding for Specimen Orientation During Embedding in Methacrylates

Polyester fibers are not dissolved by either glycol methacrylate or methyl methacrylate. Commercial polyester wadding is consequently an advantageous material to use in getting precise orientation of tissue specimens during embedding in methacrylate.     

Permanent Preservation of Whole Alizarin Red S Skeletons by Clearing and Embedding in Polyester Resins

A one-step clearing and embedding procedure for alizarin red S stained skeletons is described. Embryos are fixed in formalin, skinned and eviscerated. After staining in a 10 mg/liter solution of alizarin red S in 5% aqueous KOH, specimens are dehydrated in a graded series of acetone-polyester monomer solutions. Finally, the specimens are embedded at room temperature in the polyester resin. A special reusable metallic mold is described for embedment of large fetuses. Specimens previously cleared in glycerol can be processed with this method.     

The vascularization of the menisci. Morphological basis for the repair

Degeneration of the knee joint and increase of anterior-posterior tibial displacement are resulting from total meniscectomy, especially in knees with anterior cruciate deficiency. Vascularisation of the meniscus was studied in 12 cadaver knees after latex injection of vessels. Vascularisation of the anterior and posterior horns was found to be much better than that of the body of meniscus. All vessels originated from the popliteal artery, but variably in importance, and formed the perimeniscal and subsynovial network. 11 peripheral meniscus tears (8 freshly injured, 3 ruptures older than 2 months) were repaired by refixation, followed by immobilisation for 6 weeks. Arthroscopy 3 months postoperatively showed complete healing of all tears. Clinical examinations--follow-up for 14 months--showed no signs of rerupture in any of the patients.     

Platelet-activating factor contracts human myometrium in vitro

The myometrial contractile responses to synthetic 1-0-octadecyl-2-acetyl-sn-glyceryl-3-phosphorylcholine (platelet-activating factor, PAF) and to oxytocin were evaluated in vitro on uterine (lower segment) strips obtained from pregnant women at term (39th week), undergoing elective cesarean section. Contractility was measured isometrically in an isolated organ bath using a superfusion technique. PAF in a concentration range between 5 and 100 nM as well as oxytocin (0.1-10 mU/ml) induced a dose-dependent contraction which could be categorized in two patterns, depending on whether spontaneous activity was present. In resting strips, oxytocin induced a prompt (0.5-1 min) development of active tension, followed by a prolonged (6-18 min), slow contraction and a final relaxation. However, at variance with oxytocin, PAF-induced contractions were rhythmic (3-8/hr), and characterized by a prompt (0.5-2 min) development of tension, followed by a brief (0.5-2 min) plateau, and a final, rapid relaxation. In spontaneously active strips, both stimuli induced a marked potentiation of the contractile activity. PAF response was dependent on both cyclooxygenase- and lipoxygenase-derived products as inferred from the abrogating effects of indomethacin and FPL 55712. A receptor-mediated mechanism of action was inferred from the occurrence of specific desensitization to PAF (but not to oxytocin), and from the blocking effect of CV 3988, a specific PAF receptor antagonist. The present study indicates that PAF stimulates the contraction of human myometrium in vitro and suggests that this mediator may have a role in labor.     

Permanent preservation of whole alizarin red S skeletons by clearing and embedding in polyester resins

A one-step clearing and embedding procedure for alizarin red S stained skeletons is described. Embryos are fixed in formalin, skinned and eviscerated. After staining in a 10 mg/liter solution of alizarin red S in 5% aqueous KOH, specimens are dehydrated in a graded series of acetone-polyester monomer solutions. Finally, the specimens are embedded at room temperature in the polyester resin. A special reusable metallic mold is described for embedment of large fetuses. Specimens previously cleared in glycerol can be processed with this method.     

Effect of human T-cell leukemia virus type I tax protein on activation of the human vimentin gene.

We report that the expression of the vimentin gene, a cytoskeletal growth-regulated gene, is activated in trans by the Tax (p40x) transactivator protein encoded by the human T-cell leukemia virus type I. Expression of the Tax protein activates a number of cellular genes, such as those coding for the alpha chain of the high-affinity interleukin-2 receptor and interleukin-2. These findings indicate that the Tax protein is involved in the unregulated T-cell growth associated with human T-cell leukemia virus type I infection. Higher levels of vimentin mRNA were expressed in two human T-cell leukemia virus type I-transformed T cell lines, C91/PL and C81-66/45, when compared with that in Jurkat T cells. We demonstrate that this activation is conferred by the vimentin upstream flanking sequences. Indeed, enhanced activity was detected when constructs with the vimentin promoter linked to the chloramphenicol acetyltransferase gene were transfected in HeLa cells and in two cell lines of hematopoietic origin (Jurkat T lymphoblastoid cells and U937 promonocytic cells) together with a Tax expression plasmid. By introducing a series of deletions in the vimentin promoter, we further restrict these sequences to 30 base pairs, located between 241 and 210 base pairs upstream of the mRNA cap site. A 40-base-pair oligonucleotide containing this regulatory region proved sufficient to confer Tax inducibility upon a heterologous promoter linked to chloramphenicol acetyltransferase. Importantly, this segment includes an 11-base-pair promoter segment that has homology with the binding site for the NF-kappa B transactivating factor. Our findings indicate that constitutive expression of the vimentin gene under the control of the Tax protein may be relevant in understanding the progression of the lymphoproliferative process associated with human T-cell leukemia virus type I infection.     

Differences in diet and behaviour of sympatric saithe and pollack in a Scottish sea loch

Two closely related and morphologically similar gadoid predators, saithe, Pollachius virens and pollack P. pollachius , coexist in close proximity on a submerged reef in Loch Ewe, Scotland. The degree of overlap between the niches of these two gadoids in the wild was investigated by means of acoustic tracking, underwater television and an examination of stomach contents. Simultaneous tracking of individuals of both species revealed that pollack generally swim more slowly than saithe, restricting much of their movements to the submerged reef. Saithe ranged more widely around the reef as part of a school during the day, moving onto the reef at night. Video recordings showed that saithe swam actively and foraged in small groups and took prey items from the kelp, whereas pollack tended to remain solitary, maintained station at particular locations for minutes at a time and apparently used the kelp forest exclusively for cover. Although the dietary overlap of the two predators was considerable, their intake of different prey groups varied. In particular, within the crustaceans, saithe took amphipods, while pollack took mysids. In addition, saithe consume a wider range of prey than pollack. The relationship between the movement patterns and the use of food resources by these two predators is discussed, with particular emphasis on differences in feeding strategies.     

Effect of ethanol on membrane fluidity of protoplasts fromSaccharomyces cerevisiae andKloeckera apiculata grown with or without ethanol,measured by fluorescence anisotropy

Summary Direct measurements of membrane fluidity by fluorescence anisotropy of protoplasts fromKloeckera apiculata andSaccharomyces cerevisiae, a low and a high ethanol tolerant strain respectively, are presented. The comparison of the behaviour of the two strains grown with or without ethanol enabled us to demonstrate the existing relationship between ethanol tolerance and membrane fluidity.