Intercellular localization of nitrate reductase in roots

Experiments were conducted with segments of corn roots to investigate whether nitrate reductase (NR) is compartmentalized in particular groups of cells that collectively form the root symplastic pathway. A microsurgical technique was used to separate cells of the epidermis, of the cortex, and of the stele. The presence of NR was determined using in vitro and enzyme-linked immunosorbent assays. In roots exposed to 0.2 millimolar NO₃⁻ for 20 hours, NR was detected almost exclusively in epidermal cells, even though substantial amounts of NO₃⁻ likely were being transported through cortical and steler cells during transit to the vascular system. Although NR was present in all cell groups of roots exposed to 20.0 millimolar NO₃⁻, the majority of the NR still was contained in epidermal cells. The results are consistent with previous observations indicating that limited reduction of endogenous NO₃⁻ occurs during uptake and reduction of exogenous NO₃⁻. Several mechanisms are advanced to account for the restricted capacity of cortical and stelar cells to induce NR and reduce NO₃⁻. It is postulated that (a) the biochemical system involved in the induction of NR in the cortex and stele is relatively insensitive to the presence of NO₃⁻, (b) the receptor for the NR induction response and the NR protein are associated with cell plasmalemmae and little NO₃⁻ is taken up by cells of the cortex and stele, and/or (c) NO₃⁻ is compartmentalized during transport through the symplasm, which limits exposure for induction of NR and NO₃⁻ reduction.     

The novel flightless-I gene brings together two gene families, actin- binding proteins related to gelsolin and leucine-rich-repeat proteins involved in Ras signal transduction

The Drosophila melanogaster gene flightless-I, involved in gastrulation and muscle degeneration, has Caenorhabditis elegans and human homologues. In these highly conserved genes, two previously known gene families have been brought together, families encoding the actin- binding proteins related to gelsolin and the leucine-rich-repeat (LRR) group of proteins involved in protein-protein interactions. Both these gene families exhibit characteristics of molecular changes involving replication slippage and exon shuffling. Phylogenetic analyses of 19 amino acid sequences of 6 related protein types indicate that actin- associated proteins related to gelsolin are monophyletic to a common ancestor and include flightless proteins. Conversely, comparison of 24 amino acid sequences of LRR proteins including the flightless proteins indicates that flightless proteins are members of a structurally related subgroup. Included in the flightless cluster are human and mouse rsp-1 proteins involved in suppressing v-Ras transformation of cells and the membrane-associated yeast (Saccharomyces cerevisae) adenylate cyclase whose analogous LRRs are required for interaction with Ras proteins. There is a strong possibility that ligands for this group could be related and that flightless may have a similar role in Ras signal transduction. It is hypothesized that an ancestral monomeric gelsolin precursor protein has undergone at least four independent gene reorganization events to account for the structural diversity of the extant family of gelsolin-related proteins and that gene duplication and exon shuffling events occurred prior to or at the beginning of multicellular life, resulting in the evolution of some members of the family soon after the appearance of actin-type proteins.      

Pflanzenvergesellschaftungen mit Riccia-Arten in Südwestafrika (Namibia)

Three associations of the Nanocyperion teneriffae Lebrun 1947 (Sporoboletalia festivi Lebrun 1947) are described. They are physiognomically and ecologically similar to associations of the Mediterranean Isoeto-Nanojuncetea Br.-Bl. et Tx. 1943 and Helianthemetalia guttati Br.-Bl. 1940. They are rich in very specialized therophytes and resurrection plants.

Meinem Lehrer H. Walter, Stuttgart gewidmet.     

A 135-kd membrane protein of intercellular adherens junctions.

We report here on a new 135-kd membrane protein which is specifically associated with intercellular adherens-type junctions. This surface component was identified by a monoclonal antibody, ID-7.2.3, raised against detergent-extracted components of membranes of chicken cardiac muscle rich in intercalated discs. The antibodies stain extensively adherens junctions in intact cardiac muscle and in lens, as well as in cultured cells derived from these tissues. In living cultured cells only very little immunolabelling was obtained with ID-7.2.3 antibodies, probably due to the limited accessibility of the antibodies to the intercellular gap. However, upon the removal of extracellular Ca2+ ions a dissociation of the junction occurred, leading to the rapid exposure of the 135-kd protein. Immunoelectron microscopic labelling of EGTA-treated, or detergent-permeabilized cells indicated that the antigen is found along the plasma membrane and highly enriched in contact areas. Double immunolabelling for both the 135-kd protein and vinculin pointed to the close association of the two in intercellular junctions and to the apparent absence of the former protein from the vinculin-rich focal contacts of cultured cells and from dense plaque of smooth muscle. Immunoblotting indicated that the 135-kd protein is present in many tissues but is particularly enriched in heart, lens and brain.     

Magnesium-induced inner membrane aggregation in heart mitochondria: prevention and reversal by carboxyatractyloside and bongkrekic acid

Mg(2+) at an optimal concentration of 2mM (ph 6.5) induces large increases (up to 30 percent) in the optical density of bovine heart mitochondria incubated under conditions of low ionic strength (< approx. 0.01). The increases are associated with aggregation (sticking together) of the inner membranes and are little affected by changes in the energy status of the mitochondria. Virtually all of a number of other polyvalent cations tested and Ag(+) induce increases in mitochondrial optical density similar to those induced by Mg(2+), their approximate order of concentration effectiveness in respect to Mg(2+) being: La(3+) > Pb(2+) = Cu(2+) > Cd(2+) > Zn(2+) > Ag(+) > Mn(2+) > Ca(2+) > Mg(2+). With the exception of Mg(2+), all of these cations appear to induce swelling of the mitochondria concomitant with inner membrane aggregation. The inhibitors of the adenine nucleotide transport reaction carboxyatratyloside and bongkrekic acid are capable of preventing and reversing Mg(2+)-induced aggregation at the same low concentration required for complete inhibition of phosphorylating respiration, suggesting that they inhibit the aggregation by binding to the adenine nucleotide carrier. The findings are interpreted to indicate (a) that the inner mitochondrial membrane is normally prevented from aggregating by virtue of its net negative outer surface change, (b) that the cations induce the membrane to aggregate by binding at its outer surface, decreasing the net negative charge, and (c) that carboxyatractyloside and bongkrekic acid inhibit the aggregation by binding to the outer surface of the membrane, increasing the net negative charge.     

Differential Inhibition by Ferulic Acid of Nitrate and Ammonium Uptake in Zea mays L

The influence of the allelopathic compound ferulic acid (FA) on nitrogen uptake from solutions containing both NO₃⁻ and NH₄⁺ was examined in 8-day-old nitrogen-depleted corn (Zea mays L.) seedlings. Concurrent effects on uptake of Cl⁻ and K⁺ also were assessed. The presence of 250 micromolar FA inhibited the initial (0-1 hours) rate of NO₃⁻ uptake and also prevented development of the NO₃⁻-inducible accelerated rate. The pattern of recovery when FA was removed was interpreted as indicating a rapid relief of FA-restricted NO₃⁻ uptake activity, followed by a reinitiation of the induction of that activity. No inhibition of NO₃⁻ reduction was detected. Ammonium uptake was less sensitive than NO₃⁻ uptake to inhibition by FA. An inhibition of Cl⁻ uptake occurred as induction of the NO₃⁻ transport system developed in the absence of FA. Alterations of Cl⁻ uptake in the presence of FA were, therefore, a result of a beneficial effect, because NO₃⁻ uptake was restricted, and a direct inhibitory effect. The presence of FA increased the initial net K⁺ loss from the roots during exposure to the low K, ammonium nitrate uptake solution and delayed the recovery to positive net uptake, but it did not alter the general pattern of the response. The implications of the observations are discussed for growth of plants under natural conditions and cultural practices that foster periodic accumulation of allelopathic substances.     

Studies on the 4-carbon precursor in the biosynthesis of riboflavin. Purification and properties of L-3,4-dihydroxy-2-butanone-4-phosphate synthase

The formation of the riboflavin precursor, 6,7-dimethyl-8-ribityllumazine, from 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione requires a phosphorylated 4-carbon intermediate which has been designated as Compound X (Neuberger, G., and Bacher, A. (1985) Biochem. Biophys. Res. Commun. 127, 175-181). The enzyme catalyzing the formation of Compound X has been purified about 600-fold from the cell extract of the flavinogenic yeast Candida guilliermondii by chromatographic procedures. The purified protein appeared homogeneous as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and consisted of a single polypeptide of 24 kDa. The committed substrate of the enzyme was identified as D-ribulose 5-phosphate. The enzyme yields two products which were identified as L-3,4-dihydroxy-2-butanone 4-phosphate and formate by NMR and CD spectroscopy. Mg2+ is required for activity.