A new approach for using cofactor dependent enzymes: example of alcohol dehydrogenase

The use of enzymes requiring a cofactor as substrate in organic synthesis is still a problem since the cofactors are expensive. This study deals with a new approach consisting of using fragments of NAD+. Three fragments of NAD(H) are examined. The activities of NMN+ and NMNH are greatly improved by the addition of adenosine in ethanol oxidation and in cyclohexanone reduction, respectively. Nicotinamide mononucleoside is not active in the ethanol oxidation but the addition of AMP promotes this reaction.     

Mapping of transfer RNA genes on tobacco chloroplast DNA

Tobacco chloroplast tRNAs have been purified by two-dimensional polyacrylamide gel electrophoresis, identified by aminoacylation, labelled at their 3-end and hybridized to tobacco chloroplast DNA restriction fragments, in order to establish a tRNA gene map. These hybridization studies have revealed the localization of at least seven genes in each inverted repeat region, a minimum of 22 tRNA genes in the large single copy region and one tRNA gene in the small single copy region. Comparison of the tobacco chloroplast tRNA gene map to that of maize shows many similarities, but also some differences suggesting that DNA sequence rearrangements have occurred in the chloroplast genome during evolution.     

The xylanase introns from Cryptococcus albidus are accurately spliced in transgenic tobacco plants

The xylanase gene from Cryptococcus albidus contains seven introns. Genomic and cDNA clones under the control of the CaMV 35S promoter were transferred into tobacco plants using Agrobacterium-mediated cell transformation. The genes were transcribed and the mRNAs were amplified by the polymerase chain reaction using primers on each side of the intron region. About 90% of the amplification products from plants transformed with the genomic clone corresponded to the size of the pre-mRNA (1.2 kb) and 10% represented the spliced product (0.85 kb). The 0.85 kb fragment was cloned and sequenced and the result indicated that the introns from the xylanase gene were accurately spliced by the plant cells.     

Hydraulic architecture and water flow in growing grass tillers (Festuca arundinacea Schreb.)

The water relations and hydraulic architecture of growing grass tillers (Festuca arundinacea Schreb.) are reported. Evaporative flux density, E (mmol s^?1 m^?2), of individual leaf blades was measured gravimetrically by covering or excision of entire leaf blades. Values of E were similar for mature and elongating leaf blades, averaging 2·4 mmol s^?1 m^?2. Measured axial hydraulic conductivity, K_h (mmol s^?1 mm MPa^?1), of excised leaf segments was three times lower than theoretical hydraulic conductivity (K_t) calculated using the Poiseuille equation and measurements of vessel number and diameter. K_t was corrected (K_t*) to account for the discrepancy between K_h and K_t and for immature xylem in the basal expanding region of elongating leaves. From base to tip of mature leaves the pattern of K_t* was bell‐shaped with a maximum near the sheath–blade joint (≈ 19 mmol s^?1 mm MPa^?1). In elongating leaves, immature xylem in the basal growing region led to a much lower K_t*. As the first metaxylem matured, K_t* increased by 10‐fold. The hydraulic conductances of the whole root system, (mmol s^?1 MPa^?1) and leaf blades, (mmol s^?1 MPa^?1) were measured by a vacuum induced water flow technique. and were linearly related to the leaf area downstream. Approximately 65% of the resistance to water flow within the plant resided in the leaf blade. An electric‐analogue computer model was used to calculate the leaf blade area‐specific radial hydraulic conductivity, (mmol s^?1 m^?2 MPa^?1), using , K_t* and water flux values. values decreased with leaf age, from 21·2 mmol s^?1 m^?2 MPa^?1 in rapidly elongating leaf to 7·2 mmol s^?1 m^?2 MPa^?1 in mature leaf. Comparison of and values showed that ≈ 90% of the resistance to water flow within the blades resided in the liquid extra‐vascular path. The same algorithm was then used to compute the xylem and extravascular water potential drop along the liquid water path in the plant under steady state conditions. Predicted and measured water potentials matched well. The hydraulic design of the mature leaf resulted in low and quite constant xylem water potential gradient (≈ 0·3 MPa m^?1) throughout the plant. Much of the water potential drop within mature leaves occurred within a tenth of millimetre in the blade, between the xylem vessels and the site of water evaporation within the mesophyll. In elongating leaves, the low K_t* in the basal growth zone dramatically increased the local xylem water potential gradient (≈ 2·0 MPa m^?1) there. In the leaf elongation zone the growth‐induced water potential difference was ≈ 0·2 MPa.     

RFX proteins, a novel family of DNA binding proteins conserved in the eukaryotic kingdom.

Until recently, the RFX family of DNA binding proteins consisted exclusively of four mammalian members (RFX1-RFX4) characterized by a novel highly conserved DNA binding domain. Strong conservation of this DNA binding domain precluded a precise definition of the motif required for DNA binding. In addition, the biological systems in which these RFX proteins are implicated remained obscure. The recent identification of four new RFX genes has now shed light on the evolutionary conservation of the RFX family, contributed greatly to a detailed characterization of the RFX DNA binding motif, and provided clear evidence for the function of some of the RFX proteins. RFX proteins have been conserved throughout evolution in a wide variety of species, including Saccharomyces cerevisiae, Schizosaccharomyces pombe, Caenorhabditis elegans, mouse and man. The characteristic RFX DNA binding motif has been recruited into otherwise very divergent regulatory factors functioning in a diverse spectrum of unrelated systems, including regulation of the mitotic cell cycle in fission yeast, the control of the immune response in mammals, and infection by human hepatitis B virus.     

HGF/SF Induces Mesothelial Cell Migration and Proliferation by Autocrine and Paracrine Pathways

Mesothelial repair differs from that of other epithelial-like surfaces as healing does not occur solely by centripetal in-growth of cells as a sheet from the wound margins. Mesothelial cells lose their cell-cell junctions, divide, and adopt a fibroblast-like morphology while scattering across and covering the wound surface. These features are consistent with a cellular response to hepatocyte growth factor/scatter factor (HGF/SF). In this study, we examined the ability of mesothelial cells to secrete HGF/SF and investigated its possible role as an autocrine regulator of mesothelial cell motility and proliferation. We found that human primary mesothelial cells expressed HGF/SF mRNA and secreted active HGF/SF into conditioned medium as determined by ELISA and in a scattering bioassay. These cells also expressed the HGF/SF receptor, Met, as shown by RT-PCR and by Western blot analysis and immunofluorescence. Incubation of mesothelial cells with neutralizing antibodies to HGF/SF decreased cell migration to 25% of controls, whereas addition of HGF/SF disrupted cell-cell junctions and induced scattering and enhanced mesothelial cell migration. Furthermore, HGF/SF showed a small but significant mitogenic effect on all mesothelial cell lines examined. In conclusion, HGF/SF is produced by mesothelial cells and induces both motility and proliferation of these cells. These data are consistent with HGF/SF playing an autocrine role in mesothelial healing.     

Up regulation of corticotrophin receptors by ACTH1-24 in normal and hypophysectomized rabbits

The number of ACTH binding sites, in adrenal membranes from adult female rabbits, has been measured at different times after hypophysectomy and after ACTH_1–24 treatment. The receptor number was significantly reduced 192 h after removal of the pituitary gland as compared to intact controls. Conversely, ACTH treatment of intact rabbits enhanced the number of ACTH binding sites, or restored these levels to presurgical values in hypophysectomized animals. These results suggest that ACTH, like other hormones, is able to induce an increase in the number of its own receptors; the physiological significance of such variations remains however to be elucidated.