Leghemoglobin in Lupin Plants (Lupinus albus cv Multolupa)

Leghemoglobin was localized by immunogold techniques in nodules of Lupinus albus cv Multolupa inoculated with Bradyrhizobium sp. (Lupinus) strain ISLU 16. The protein localization was performed in nodules embedded in Spurr's and Araldite epoxy resins and Lowycryl K4M. A very good preservation of both the ultrastructure and antigenicity was obtained with the tissues embedded in Araldite following glutaraldehyde fixation and unpostfixed in osmium tetroxide. Lupin leghemoglobin is a stable and abundant protein which allows a conventional method to be safely used for localization of leghemoglobin. Labeling of leghemoglobin was specifically confined to the cytosol matrix and nuclei. Gold particles were never observed in the peribacteroidal spaces nor in the cytoplasmic organelles of the infected cells. Decrease of leghemoglobin was observed when the plants were grown with 10.7 micromolar and 21.4 micromolar of nitrate.     

Evaluation of strain competitiveness in Rhizobium leguminosarum bv phaseoli using a nod + fix− natural mutant

Competition from native soil rhizobia is likely to be an important factor limiting Phaseolus vulgaris L. inoculant response in Latin America. We used UMR 1116, a nod ⁺ fix^– natural mutant of Rhizobium leguminosarum bv phaseoli strain CC511, as a reference strain to study competition for nodulation sites in this species. When P. vulgaris cv Carioca was planted in soils containing different proportions of UMR 1116 and the effective and competitive strain UMR 1899, UMR 1116 occupied more than 50% of the nodules at all inoculant ratios tested, though increasing the proportion of UMR 1899 in the inoculant did enhance the number and percentage of effective nodules and plant dry weight. Sixty two strains of bean rhizobia were tested in competition with UMR 1116. An inoculant ratio of 1:1 was used, with all strains applied to the soil rather than to seeds. Strains varied in the number and percentage of effective nodules produced in competition with UMR 1116, and in plant dry weight, and there was a strong correlation between variation in each of these traits and plant N accumulation. Seven of the strains (UMR 1073, 1084, 1102, 1125, 1165, 1378 and 1384) were identified as both superior in competitive ability and active in N₂ fixation. Site of placement of the inoculant and ambient temperature influenced strain response.Journal paper 16736, Agricultural Experiment Station, University of Minnesota, St. Paul, MN 55108, USA     

Mechanism of action of levonorgestrel: in vitro metabolism and specific interactions with steroid receptors in target organs.

Levonorgestrel (LNG) is a synthetic steroid that displays potent progestional and androgenic effects but it lacks estrogen-like activity. To examine the mode of action of this progestin, we studied its metabolism in vitro in target organs and the specific interactions of LNG and its metabolites with putative steroid receptors. The results demonstrated that [³H]LNG was efficiently converted to A-ring reduced derivatives when incubated with rat hypothalamus and pituitary. Under optimal incubation conditions, [³H]5-dihydro LNG (5-LNG) and [³H]3-5-tetrahydro LNG (3,5-LNG) were identified as the major metabolic conversion products, while [³H]3ß, 5-LNG formation occured to a lesser extent. A-ring reduction of LNG was NADPH-dependent. Assessment of the relative binding affinities of LNG and its derivatives to progesterone (PR), androgen (AR) and estrogen (ER) receptors by displacement analysis revealed that unchanged LNG binds with high affinity to PR and AR but not to ER. 5-LNG exhibited a diminished though significant interactions with PR and an enhanced binding affinity for AR as compared with LNG, indicating that 5-reduction of LNG increases its affinity for AR. The most striking finding was that further reduction of the 5-LNG molecule at C-3 abolished its binding activity to PR, AR, and even to ER. The overall data provides a plausible explanation for the lack of estrogen agonistic action of LNG and for its potent progestational and androgenic effects.     

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.     

Mapping in the mouse of the region homologous to the rat growth and reproduction complex (grc)

Offprint requests: J. Klein.     

Properties of Chicken Lens MIP Channels Reconstituted into Planar Lipid Bilayers

Membrane fractions highly enriched in chicken lens MIP (MIP28) were found to form ion channels when incorporated into planar lipid bilayers. The channels displayed prominent unitary conductances of about 60 and 290 pS in symmetric 150 mm KCl solution and were slightly anion selective. For both depolarizing and hyperpolarizing voltages, voltage sensitivity of the MIP28-induced conductance could be fit by a Boltzmann relation, symmetric around zero mV, with V ₀ = 18.5 mV, n= 4.5 and g _min/g _max= 0.17. Channel properties were not appreciably altered by pH in the range of 5.8 to 7, although channel incorporation was observed to occur more frequently at lower pH values. Calcium, at millimolar concentrations, decreased the channel mean open time. Partial proteolysis of MIP28 to yield MIP21 did not appreciably affect single-channel conductance or voltage sensitivity of the reconstituted channels. MIP28 was not phosphorylated by cAMP dependent protein kinase (PKA). Although unitary conductance and selectivity of the chicken MIP channel are similar to those reported for the bovine MIP (MIP26), the voltage sensitivity of MIP28 was higher than that of the bovine homologue, and voltage sensitivity of MIP28 was not modulated by treatments previously shown to affect MIP26 voltage gating (partial proteolysis and protein phosphorylation by PKA: (Ehring et al., 1990). The existence of such strikingly different functional properties in highly homologous channel isoforms may provide a useful system for exploration of the structure-function relations of MIP channels. Received: 27 March 1996/Revised: 5 August 1996     

Indirect evidence of alteration in the expression of the rDNA genes in interspecific hybrids betweenDrosophila melanogaster andDrosophila simulans

Crosses betweenDrosophila melanogaster females andD. simulans males produce viable hybrid females, while males are lethal. These males are rescued if they carry theD. simulans Lhr gene. This paper reports that females of the wild-typeD. melanogaster population Staket do not produce viable hybrid males when crossed withD. simulans Lhr males, a phenomenon which we designate as the Staket phenotype. The agent responsible for this phenomenon was found to be the StaketX chromosome (X ^mel ,Stk). Analysis of the Staket phenotype showed that it is suppressed by extra copies ofD. melanogaster rDNA genes and that theX ^mel ,Stk chromosome manifests a weak bobbed phenotype inD. melanogaster X ^mel ,Stk/0 males. The numbers of functional rDNA genes inX ^mel ,Stk andX ^mel ,y w (control) chromosomes were found not to differ significantly. Thus a reduction in rDNA gene number cannot account for the weak bobbedX ^mel ,Stk phenotype let alone the Staket phenotype. The rRNA precursor molecules transcribed from theX ^mel ,Stk rDNA genes seem to be correctly processed in both intraspecific (melanogaster) and interspecific (melanogaster-simulans) conditions. It is therefore suggested that theX ^mel ,Stk rDNA genes are inefficiently transcribed in themelanogaster-simulans hybrids.     

Stimulation of in vitro shoot proliferation from nodal explants of cassava by thidiazuron, benzyladenine and gibberellic acid

Multiple shoots were produced from nodal explants of cassava (Manihot esculenta Crantz) by a two-step procedure: a 6- to 8-day exposure to 0.11–0.22 µM thidiazuron (TDZ) in liquid Murashige and Skoog (MS) medium followed by culture on agar-solidified MS medium supplemented with 2.2 µM 6-benzyladenine (BA) and 1.6 M gibberellic acid (GA₃). TDZ caused the nodal explants to expand and this expansion (growth) continued during culture with BA and GA₃. From this expanded explant, clusters of buds and fasciated stems developed continuously and these gave rise to shoots. The shoot proliferation process was open-ended, yielding an average of 31.5 shoots per nodal explant after 10 weeks of culture with genotype CG 1–56. A positive response was also obtained from seven other genotypes evaluated with this protocol.Abbreviations BA 6-benzyladenine - BM basal medium - DPU 1,3-diphenylurea - GA₃ gibberellie acid - 2iP isopentenyladenine - MSM multiple shoot medium - NAA 1-naphthaleneacetic acid - PGR plant growth regulator - TDZ thidiazuron - Z zeatin     

Paracoccidioides brasiliensis antigen batches from the same isolate show immunological and biochemical differences

We investigated the occurrence of antigenic and biochemical variability among Paracoccidioides brasiliensis antigen batches prepared according to the same protocol. Initially (experiment #1), we analyzed two antigen lots of two human isolates (Bt1 & Bt2), cultured in two media (PYG: bactopeptone, yeast extract, glucose; MMM: McVeigh & Morton medium) in SDS-PAGE and in two immunological tests (imunodiffusion-ID and footpad swelling test-FPT). Afterwards (experiment #2), we compared the antigenic profile of three antigen batches from three human isolates (Bt1, Bt2 & Bt3) by two-dimensional immunoelectrophoresis (2 D-IEP) against a reference system for P. brasiliensis antigens. In experiment #1, there were important intra- and inter-strain antigenic differences between batches of the fungal isolates cultured on both media. The block titration of the antigen batches for the immunological tests revealed correlation between protein concentration and biological activity in ID and no correlation in FPT. In experiment #2, the reference system for P. brasiliensis showed 26 antigen peaks. There were important differences between batches prepared from the same isolate and between batches from different isolates. Our data suggested the occurrence of instability in the synthesis of antigenic components by a same P. brasiliensis isolate, under controlled incubation conditions.