Developmental regulation of presence of binding sites for neoglycoproteins and endogenous lectins in various embryonic stages of human lung,liver and heart

Protein-carbohydrate interactions are supposed to play key roles in the mechanisms of cell adhesion, biosignalling and intracellular routing, warranting the analysis of the developmental course of expression of epitopes of this system. Thus, a panel of carrier-immobilized carbohydrate ligands was used as probes, namely lactose,N-acetylgalactosamine,N-acetylglucosamine, mannose, fucose and maltose. Additionally, an antibody to an endogenous -galactoside-binding lectin (anti-galectin-1), the biotinylated lectin and two further human lectins, namely the macrophage migration inhibitory factor-binding sarcolectin and serum amyloid P component (SAP) that displays selectivity for sulphated sugars and mannose-6-phosphate, were included. They enabled us to assess the extent of the presence of respective binding sites in fixed sections from human lungs (pulmonary epithelial cells), livers (hepatocytes) and hearts (myocard cells) of 10–50 weeks gestation. Invariably, specific binding was detected in the three organ types, at least in certain stages. In most of the cases, the intensity of staining exhibited developmental regulation. The apparent patterns reveal similarities between the different cell types, as seen with immobilizedN-acetylglucosamine as well as with labelled galectin-1 and sarcolectin. However, drastic differences among such patterns with nearly opposite developmental courses do also occur, as detected for carrier-attached mannose and maltose residues. These results point to a potential importance for the detected glycohistochemical features in human development and substantiate the possibility of differential regulation of the presence of binding sites for distinct sugars within a certain organ and between the individual cell types of the monitored organs.     

Genetic analysis of the yeastCandida maltosa by means of induced parasexual processes

The usefulness of hybridization by protoplast fusion and mitotic segregation for the genetic analysis of the imperfect fodder yeastCandida maltosa was tested. Mitotically stable fusion hybrids were obtained with frequencies between 10^–6 and 10^–7. Complementation tests were performed by protoplast fusion. Substances that are known to induce frequent mitotic segregation in other yeast species such as benomyl, p-fluorophenylalanine, and acriflavine were ineffective inC. maltosa. UV irradiation induced mitotic segregation in up to 10%. This agent induced mainly mitotic crossing over inC. maltosa. Our data enabled the construction of the linkage group I with the sequenceCEN-ade-26-pro-1.     

The conversion of [3H] tryptophan to 5-[3H] hydroxytryptamine in mouse brain following depletion of phenylalanine and tyrosine.

Abstract— Phenylalanine ammmonia-lyase (PAL), an enzyme which converts phenylalanine (Phe) and tyrosine (Tyr) to trans-p-cinnamic acid and trans-p-coumaric acid, respectively, was administered to mice and its effect on the conversion of [³H]tryptophan to 5-[³H]HT in the brain was measured. Although PAL significantly depleted plasma Tyr, it has little or no effect on either brain Tyr or catecholamine concentrations. Endogenous brain tryptophan levels were significantly increased 2 h after PAL administration, brain 5-HT was dramatically increased 4 h following PAL and each returned to baseline levels by 8 h. This return to baseline was accompanied by a marked decrease in the fraction of tryptophan converted to 5-HT during a 20 min pulse period preceding death, suggesting the activation of a compensatory decrease in 5-HT synthesis in response to increased 5-HT concentration. These data suggest that PAL administration readily produces reversible alterations in 5-HT synthesis and that this may be a fruitful approach to studying brain 5-HT function.     

Integration specificity of an artificial kanamycin transposon constructed by the in vitro insertion of an internal Tn5 fragment into IS2

Molecular Genetics and Genomics - IS2 has been marked genetically by the in vitro insertion into its HindIII site of a 3.3 Kb HindIII fragment of Tn5 conferring resistance to kanamycin. The...     

Chemical and Physical Characterization of Interfacial-Active Lipids from Rhodococcus erythropolis Grown on n-Alkanes

Lipophilic compounds of the culture suspension containing Rhodococcus erythropolis DSM43215 had surfactant properties when the bacteria were cultivated with n-alkanes as the sole carbon source. Thirteen main components from a dichloromethane-methanol extract of the R. erythropolis cultures were isolated and characterized to specify quantitatively their surfactant properties, e.g., minimum surface and interfacial tensions and critical micelle concentrations. The interfacial activity of the organic extract was dominated by α,α-trehalose-6,6′-dicorynomycolates which reduced interfacial tension from 44 to 18 mN/m. Phosphatidylethanolamines which were also present in the organic extract reduced the interfacial tension below 1 mN/m. The trehalose corynomycolates had extremely low critical micelle concentrations in high-salinity solutions, and the interfacial properties were stabile in solutions with a wide range of pH and ionic strength.     

Transforming growth factor-beta and platelet-derived growth factor synergistically stimulate contraction by testicular peritubular cells in culture in serum-free medium

We report investigations on factors influencing contractility by testicular peritubular cells (PC) maintained in culture in a three-dimensional collagen gel system, and the behavior of PC in culture on a two-dimensional system. At low and moderate cell densities, PC embedded in collagen gels in serum-free Eagle's minimal essential medium (MEM) have a lesser degree of contractility than PC in culture in MEM containing calf serum. The contractility by PC, measured by determining changes in diameter of the collagen gel, was increased by addition of transforming growth factor-beta (TGF-beta) to serum-free MEM, and this was further enhanced by supplementing the medium with platelet-derived growth factor (PDGF). In the absence of TGF-beta, however, PDGF had no detectable effects on PC contractility. Other growth factors examined (epidermal growth factor, insulin, and fibroblast growth factor) did not influence the degree of contractility of PC in serum-free MEM in the presence or absence of TGF-beta. PC maintained in MEM supplemented with platelet-poor serum (PPS) have a lesser degree of contractility than their counterparts in MEM containing 2.5% calf serum. The addition of TGF-beta and PDGF to PPS-supplemented MEM restored contractility by PC to a level comparable to that observed by PC in MEM containing complete serum. The addition of nonpurified bovine serum albumin (BSA) to MEM greatly increased PC contractility. By contrast, highly purified BSA had no such effect, suggesting that one or more components adsorbed to the impure BSA was implicated. Polyclonal antibody against fibronectin did not influence the contractility of PC in collagen gels in the presence or absence of serum. Antiserum against TGF-beta partially blocked the enhancement of contractility of PC in MEM containing non-purified BSA. In PC plated on top of a collagen gel lattice, the attachment, spreading, and cell shape were greatly influenced by the presence of TGF-beta and PDGF, both singly and together. Data presented are interpreted to indicate that effects elicited by serum on the properties of PC in culture, and on the contractility of PC, can be attributed in part to the combined influences of TGF-beta and PDGF in serum.     

Marine biosurfactants. IV. Production,characterization and biosynthesis of an anionic glucose lipid from the marine bacterial strain MM1

Summary During screening for biosurfactants among marine, n-alkane-utilizing bacteria, low- and high-molecular surface-active substances were detected. The marine bacterial strain MM1 was found to synthesize a novel glycolipid that has not so far been cited in the literature. Both ¹H, ¹³C-nuclear magnetic resonance spectroscopic and positive ion fast atom bombardment mass spectrometer studies led to the identification of a glucose lipid consisting of four 3-OH-decanoic acids, which are linked together by ester bonds. The lipophilic moiety is coupled glycosidically with C-1 of glucose. The glucolipid reduced the surface tension from 72 mN/m to 30 mN/m while the minimum interfacial tension towards n-hexadecane was lowered to values smaller than 5 mN/m. Correspondence to: S. Lang     

Directed mutagenesis of DNA cloned in filamentous phage: influence of hemimethylated GATC sites on marker recovery from restriction fragments.

Gapped duplex DNA molecules of recombinant genomes of filamentous phage are constructed in vitro. Denatured restriction fragments covering (part of) the precisely constructed gap are hybridized to the gapped duplex DNA molecules to form ternary duplices. The two strands of the ternary duplex molecules carry different genetic markers within the region spanned by the restriction fragment leading to a one base pair mismatch or to an insertion loop of 93 nucleotides, respectively. The two strands also vary with respect to A-methylation in GATC sites. In cases of asymmetrical methylation, transfection of E. coli with these heteroduplex molecules leads to marker recoveries with a pronounced bias in favour of the marker encoded by the methylated strand. This effect at least partly explains the comparably low marker yields achieved in previous directed mutagenesis experiments using filamentous phage as the vector. The results suggest how these procedures can be optimized. Precise construction of a 93 bp insertion of 9.5% marker yield is described.