Expression cloning and regulation of steroid 5 alpha-reductase, an enzyme essential for male sexual differentiation

The conversion of testosterone into the more potent androgen, dihydrotestosterone, catalyzed by the enzyme steroid 5 alpha-reductase, is required for the differentiation of male external genitalia. Here, we report the isolation of cDNA clones encoding the rat steroid 5 alpha-reductase using expression cloning in Xenopus oocytes. DNA sequence analysis demonstrates that the liver and ventral prostate forms of steroid 5 alpha-reductases are identical hydrophobic proteins of 29 kDa. The amount of steroid 5 alpha-reductase mRNA in liver increased in response to castration, but remained unchanged in the prostate. Testosterone administration to castrates induced expression of mRNA in the prostate but had no effect on liver. The data suggest that the steroid 5 alpha-reductase gene is differentially regulated by testosterone in androgen-responsive versus non-responsive tissues.     

Biogenesis of the somatogenic receptor in rat liver

Certain structural characteristics, in particular the type of oligosaccharide chains associated with the rat liver somatogenic (GH) receptors, were studied in different isolated organelles involved in receptor biosynthesis, maturation, and binding, with the use of ligand-affinity cross-linking, incubation with various oligosaccharide chain-cleaving enzymes, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In an endoplasmic reticulum-enriched fraction, a somatogenic receptor with Mr 33,000, after correction for bound ligand (assuming a 1:1 binding ratio of ligand to receptor) was found to contain N-linked high mannose oligosaccharide chain(s). In an intermediate density fraction, enriched in cis-Golgi, a major receptor of Mr 43,000 was found to contain N-linked complex type of oligosaccharide chains. In a low density membrane fraction, containing trans-Golgi complex membranes and endocytic vesicles, three receptors of Mr 95,000, 55,000, and 43,000 were found. These three receptors contain N-linked complex-type oligosaccharide chains. Neuraminidase treatment resulted in a decrease of the Mr 95,000 and 43,000 receptors to Mr 81,000 and 39,000, respectively. Two specific somatogenic receptors of Mr 95,000 and 43,000 containing N-linked complex type of oligosaccharides were found in an isolated plasma membrane-enriched fraction. When isolated hepatocytes were analyzed, the Mr 95,000 receptor was found to be the major labeled species. Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis (first dimension nonreducing and the second dimension reducing conditions), showed that the Mr 43,000 receptor is contained within the Mr 95,000 receptor. The data suggest that the Mr 33,000 receptor found in endoplasmic reticulum constitutes a precursor to the Mr 43,000 receptor and that the Mr 43,000 receptor is complexed with an unknown subunit during transport through the Golgi complex to form an Mr 95,000 receptor present on the cell surface.     

Rotational and translational movement features of the pelvis and thorax during adult human locomotion

The kinematics of the pelvis and thorax are important in gait studies since their movement patterns are closely related to gait efficiency and 'smoothness' of locomotion. The purpose of this study was to identify features of normal gait patterns for later comparisons with pathological and developmental gait patterns. A two camera SELSPOT system interfaced with an HP1000 minicomputer was used to obtain three-dimensional kinematic/temporal data for the pelvis and thorax. Data from treadmill walking of eight adults were used for within subject (at different speeds) analyses. The analyses revealed a very complex pattern with a set of breakpoints which was consistent over all subjects. Some features were invariant over a range of walking speeds although the total range of motion changed considerably.     

Mutations induced by benzo[a]pyrene diolepoxide at the hprt locus in human T-lymphocytes in vitro

Human T-lymphocytes have been treated with benzo[a]pyrene diolepoxide (BPDE) in vitro and T-cell clones mutated in the hprt gene have been isolated. The mutant frequencies in BPDE-treated T-cell cultures were on average 24-fold higher than those of untreated cultures. Thus, BPDE is a potent inducer of gene mutation in this system. In order to examine which types of mutations are induced by BPDE in human cells, 41 spontaneous and 44 BPDE-induced mutant clones have been characterized using the Southern blot technique. In addition, rearrangements of the T-cell-receptor beta and gamma loci have been used to determine the proportion of isolated clones that are unique, and thus likely to represent independent mutational events. Out of 23 independent spontaneous mutants 4 had large hprt alterations that could be detected on Southern blots. Two of these alterations, deletions of exons 2-6, have been confirmed using PCR of hprt cDNA and direct sequencing of the PCR product. All 33 independent BPDE-induced mutants had normal hprt restriction patterns which indicates that BPDE is mainly a point mutagen in this system.     

Bioconversions in aqueous two-phase systems.

Bioconversions involving enzymes and/or microbial cells in aqueous two-phase systems are reviewed. The partitioning of biocatalysts, substrates, and products is discussed in relation to their size. The efficiency of retaining biocatalysts in aqueous two-phase systems is summarized in relation to other methods of recirculating. The influence of phase components on the activity and the stability of enzymatic biocatalysts is exemplified with penicillin acylase and the cellulolytic enzyme system, and the effect of phase components on biocatalytic living cells is exemplified with the production of alpha-amylase with Bacillus sp. Process design costs in bioconversions in aqueous two-phase systems are briefly summarized.     

Systematics of theLactobacillus population on rat intestinal mucosa with special reference toLactobacillus reuteri

The systematics of theLactobacillus population of the intestines of 88 different rats was studied; 80 rats had been fed on fermented oat-meal soup (Molin et al. 1992). One-hundred-twenty-twoLactobacillus strains from the intestinal mucosa were phenotypically classified together with twenty-eight reference strains ofLactobacillus andLeuconostoc, using 49 unit characters. Data were examined using Jaccard coefficient, and unweighted pair group algorithm with arithmetic averages. Two major and eleven minor clusters were defined at the 76% S_J-similarity level: Cluster 1 included thirty isolates which could not be identified further, but had resemblance to the type strains ofL. jensenii, L. gasseri, L. crispatus, and to some extent toL. acidophilus. Cluster 12 including fifty-four intestinal isolates was identified asL. reuteri; and so was cluster 13 (five isolates). Isolates of the major clusters were found in all parts of the intestines. The genomic homogeneity of theL. reuteri isolates was scrutinized by endonuclease restriction analysis of the chromosomal DNA, and the isolates could be divided into six genomic strains.     

Sexual selection

Competition over mates takes many forms and has far-reaching consequences for many organisms. Recent work suggests that relative reproductive rates of males and females, sperm competition and quality variation among mates affect the strength of sexual selection. Song, other display, body size, visual ornaments and material resource offerings are often sexually selected. There is much empirical evidence of mate choice, and its evolution is clarified by mathematical models. Recent advances in theory also consider costs of choice, effects of deleterious mutations, fast and slow evolution of preferences and preferred traits, and simultaneous preferences for several traits. Contests over mates are important; so is sperm competition, scrambles, endurance rivalry, and coercion. The latter mechanisms have received less attention than mate choice. Sexual selection may explain puzzling aspects of plant pollination biology.     

Immunoaffinity column clean-up for the high-performance liquid chromatographic determination of aflatoxins B1, B2, G1, G2, M1 and Q1 in urine

A method for the determination of aflatoxins B₁, B₂, G₁, G₂, M₁ and Q₁ in human urine has been developed. The 10-ml urine samples were automatically cleaned up on immunoaffinity columns and analysed by high-performance liquid chromatography (HPLC), including post-column derivatization with bromine and fluorescence detection. Average aflatoxin recoveries were: B₁ 103%, B₂ 106%, G₁ 98% and G₂ 96% in the range 6.8–73 pg/ml of urine and M₁ 103% and Q₁ 100% in the range 18–97 pg/ml of urine. The relative standard deviations were all between 1% and 21%. The determination limits of aflatoxins in urine were 6.8 pg/ml for B₁, B₂, G₁ and G₂ and 18 pg/ml for M₁ and Q₁.