Intramolecular base composition heterogeneity of human DNA.

The intramolecular base composition heterogeneity of human DNA has been investigated by electron microscopic observations of partially denatured structures and by equilibrium solution thermal denaturation techniques. DNA sequences having an average length of less than 2000 base pairs are found to be heterogeneous in base composition. These heterogeneous sequences occupy a minimum of 67 to 81% of the human genome.     

Chromosome analyses of children after ecological lead exposure

In the present work chromosome analysis was performed in a group of 30 children living in a town with a lead plant. Due to the emission of the smelter the individual lead uptake through food, drinking water and inhalation was increased. They were selected out of 1600 children whose blood lead level, delta-aminolevulinic acid dehydratase activity in the erythrocytes and erythrocyte porphyrine level was measured. In the investigated group of children the values of these parameters showed to be indicative for a significant lead exposure. A total of 10,000 cells was scored after 48 h culture time. Despite a significantly increased lead load as compared with two groups of 10 children from a suburb and the isle of Helgoland there was neither evidence for a higher number of cells with structural chromosome aberrations, nor for an increased aberration yield.     

The organization of macronuclear DNA sequences associated with C4A4 repeats in the polytene chromosomes of Stylonychia lemnae

Gene libraries of the micronucleus and the macronuclear anlagen of the polytene chromosome stage of Stylonychia lemnae were screened for internal C₄A₄ repeats. The number of these internal repeats was shown to be identical in both kinds of nuclei. Analysis of macronuclear sequences associated with C₄A₄ in the polytene chromosomes showed that several macronuclear DNA sequences are clustered. However, interspersed between short exons of one gene are located exons of several other genes, i.e. the exon of one gene is an intron for several other genes.by M. Trendelenburg     

Early ventral expression of the Drosophila neurogenic locus mastermind

The neurogenic locus mastermind (mam) of Drosophila is required for the segregation of epidermal from neural cell lineages. Previous studies have shown that during neurogenesis mam appears to be expressed throughout the ectoderm, mesoderm, and neuroblast layer of the germ band. Here it is demonstrated that during early embryogenesis mam is expressed ubiquitously; however, the predominant domains of accumulation of mam RNA and protein during gastrulation are along the ventral longitudinal surface, including cells of the mesoderm, endoderm, mesectoderm, and neuroectoderm. The regions of elevated mam accumulation coincide with the realm of action of dorsoventral patterning genes.     

Comparative study of the structure/function relationship of wild-type and structurally modified maltopentaose-producing amylase.

Amylase A-180, which is secreted by a new alkaliphilic organism, isolate 163-26, consists of a single type of polypeptide chain of 186.5 kDa and hydrolyses starch by exo-attack releasing malto-pentaose as preferential product. The structure/function relationship of this unusual starch-degrading enzyme was analysed by introducing 3' deletions into the structural gene. It was found that removal of up to a 110-kDa portion from the C-terminus leaving 563 N-terminal amino acids still led to the formation of a fully active enzyme. The part of the structural gene coding for these 563 N-terminal amino acids was fused with the signal peptide-encoding segment of the cyclodextrin glucanotransferase gene from Klebsiella oxytoca and was cloned into an expression vector. The resulting truncated A-180 derivative, A-180/21, was efficiently transported through the cytoplasmic membrane and released into the medium by an Escherichia coli strain which 'leaks' periplasmatic components. A-180/21 was purified and its catalytic properties, i.e. specific activity and product specificity, proved to be identical to those of the wild-type enzyme; however, in contrast to the wild-type enzyme, it was unable to bind to raw starch and it displayed an altered temperature and pH dependence of activity.     

Cytosol- and clathrin-dependent stimulation of endocytosis in vitro by purified adaptors

Using stage-specific assays for receptor-mediated endocytosis of transferrin (Tfn) into perforated A431 cells we show that purified adaptors stimulate coated pit assembly and ligand sequestration into deeply invaginated coated pits. Late events in endocytosis involving membrane fission and coated vesicle budding which lead to the internalization of Tfn are unaffected. AP2, plasma membrane adaptors, are active at physiological concentrations, whereas AP1, Golgi adaptors, are inactive. Adaptor-dependent stimulation of Tfn sequestration requires cytosolic clathrin, but is unaffected by clathrin purified from coated vesicles suggesting that soluble and assembled clathrin pools are functionally distinct. In addition to adaptors and cytosolic clathrin other, as yet unidentified, cytosolic factors are also required for efficient coated pit invagination. These results provide new insight into the mechanisms and regulation of coated pit assembly and invagination.     

The complete primary structure of the boar spermadhesin AQN-1, a carbohydrate-binding protein involved in fertilization.

Gamete recognition and adhesion are essential steps in the complex process of fertilization. In mammals and in other species, increasing evidence indicates that carbohydrate-binding proteins on the sperm surface play a pivotal role as counter-receptors for certain oligosaccharide moieties attached to the oocyte zona pellucida glycoproteins. Although different sperm-associated zona-pellucida-binding proteins have been identified in a number of species, few of them have been isolated and structurally characterized. In this paper we report the primary structural characterization of AQN-1, a 12-kDa boar-sperm-associated carbohydrate-binding and zona-pellucida-binding protein. The molecular mass of AQN-1 was determined by time-of-flight plasma-desorption mass spectrometry. Determination of its amino acid sequence and location of disulphide bridges were accomplished by a combination of proteochemical and mass spectrometric methods. The primary structure of AQN-1 failed to show any significant similarity to the protein structures deposited with the Martinsried Institute for Protein Sequences data bank, indicating that it may belong to a novel protein family involved in fertilization. AQN-1 shares extensive structural, as well as functional, similarity with two other boar sperm zona-pellucida-binding proteins, AQN-3 and AWN, which we have recently characterized. To name this protein family, we have coined the term spermadhesin. Our data may be relevant for identification of spermadhesins in other species, and thus may contribute to a better understanding of the species-specific sperm-egg recognition mechanism.     

Intact but not truncated insulin-like growth factor binding protein-3 (IGFBP-3) blocks IGF I-induced stimulation of osteoblasts: control of IGF signalling to bone cells by IGFBP-3-specific proteolysis?

IGFBP-3 is the predominant IGFBP in serum and the major IGFBP secreted by osteoblasts. Native and recombinant IGFBP-3 and a truncated form lacking the carboxyterminal third were tested for their effects on 2 osteoblastic cell lines. Intact but not truncated IGFBP-3 blocked IGF I-stimulated DNA and glycogen synthesis. Inhibition was dose-dependent and found whenever the concentration of intact IGFBP-3 exceeded the concentration of IGF I. Truncated IGFBP-3 appears to result from proteolytic cleavage and does occur in vivo. The loss of inhibition by IGFBP-3 may be regulated at the site of IGF target cells and thus be essential for IGF I-induced osteoblast growth.     

Effects of L-proline and post-plating temperature treatment on Maize (Zea mays L.) anther culture

Summary Comparison of different post-plating temperature regimes with a control treatment (27° C) revealed that a short-term cold (8/14°C:2/2 days or 14°C:4 days) as well as a heat treatment (30°C:14 days) increased the production of embryro-like-structures (ELS) from cultured maize anthers. The beneficial effects of short-term cold treatments were magnified 2–3 times when L-proline (PROL) was added to the induction medium (125–500 mg/L). In the best treatment (14°C:4 days, 125 mg/L L-proline) one genotype produced 143.5 ELS/100 anthers. Anthers subjected to high temperature (30°C:4 days, 30°C:7 days, 30°C:14 days) generally showed a lower response than did cold treated anthers, although genotypic differences were observed. Regeneration frequency did not appear to be affected by the presence of L-proline in the induction medium.Abbreviations ELS Embryo-like-structures - PROL L-proline