Modification of chemical properties of cell walls by silicon and its role in regulation of the cell wall extensibility in oat leaves

Effects of silicon on the mechanical and chemical properties of cell walls in the second leaf of oat (Avena sativa L.) seedlings were investigated. The cell wall extensibility in the basal region of the second leaf was considerably higher than that in the middle and subapical regions. Externally applied silicon increased the cell wall extensibility in the basal region, but it did not affect the extensibility in the middle and subapical regions. The amounts of cell wall polysaccharides and phenolic compounds, such as diferulic acid (DFA) and ferulic acid (FA), per unit length were lower in the basal region than in the middle and subapical regions of the leaf, and silicon altered these amounts in the basal region. In this region, silicon decreased the amounts of matrix polymers and cellulose per unit length and of DFA and FA, both per unit length and unit matrix polymer content. Silicon treatment also lowered the activity of phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) in the basal region. In contrast, the amount of silicon in cell walls increased in response to silicon treatment in three regions. These results suggest that in the basal region, silicon reduces the net wall mass and the formation of phenolic acid-mediated cross-linkages between wall polysaccharides. Such modifications of wall architecture may be responsible for the silicon-induced increase in the cell wall extensibility in oat leaves.     

Mechanism of rapid-phase insulin response to elevation of portal glucose concentration

The hepatoportal region is important for glucose sensing; however, the relationship between the hepatoportal glucose-sensing system and the postprandial rapid phase of the insulin response has been unclear. We examined whether a rapid-phase insulin response to low amounts of intraportal glucose infusion would occur, compared that with the response to intrajugular glucose infusion in conscious rats, and assessed whether this sensing system was associated with autonomic nerve activity. The increases in plasma glucose concentration did not differ between the two infusions at 3 min, but the rapid-phase insulin response was detected only in the intraportal infusion. A sharp and rapid insulin response was observed at 3 min after intraportal infusion of a small amount of glucose but not after intrajugular infusion. Furthermore, this insulin response was also induced by intraportal fructose infusion but not by nonmetabolizable sugars. The rapid-phase insulin response at 3 min during intraportal infusion did not differ between rats that had undergone hepatic vagotomy or chemical sympathectomy with 6-hydroxydopamine compared with control rats, but this response disappeared in rats that had undergone chemical vagotomy with atropine. We conclude that the elevation of glucose concentration in the hepatoportal region induced afferent signals from undetectable sensors and that these signals stimulate pancreas to induce the rapid-phase insulin response via cholinergic nerve action.     

Osteoprotegerin reduces the serum level of receptor activator of NF-kappaB ligand derived from osteoblasts

Osteoprotegerin (OPG) is a decoy receptor for receptor activator of NF-kappaB ligand (RANKL). We previously reported that OPG deficiency elevated the circulating level of RANKL in mice. Using OPG(-/-) mice, we investigated whether OPG is involved in the shedding of RANKL by cells expressing RANKL. Osteoblasts and activated T cells in culture released a large amount of RANKL in the absence of OPG. OPG or a soluble form of receptor activator of NF-kappaB (the receptor of RANKL) suppressed the release of RANKL from those cells. OPG- and T cell-double-deficient mice showed an elevated serum RANKL level equivalent to that of OPG(-/-) mice, indicating that circulating RANKL is mainly derived from bone. The serum level of RANKL in OPG(-/-) mice was increased by ovariectomy or administration of 1alpha,25-dihydroxyvitamin D(3). Expression of RANKL mRNA in bone, but not thymus or spleen, was increased in wild-type and OPG(-/-) mice by 1alpha,25-dihydroxyvitamin D(3). These results suggest that OPG suppresses the shedding of RANKL from osteoblasts and that the serum RANKL in OPG(-/-) mice exactly reflects the state of bone resorption.     

Epstein-Barr virus BZLF1 gene, a switch from latency to lytic infection, is expressed as an immediate-early gene after primary infection of B lymphocytes

We demonstrate here that the Epstein-Barr virus (EBV) BZLF1 gene, a switch from latent infection to lytic infection, is expressed as early as 1.5 h after EBV infection in Burkitt's lymphoma-derived, EBV-negative Akata and Daudi cells and primary B lymphocytes. Since BZLF1 mRNA is expressed even when the cells are infected with EBV in the presence of anisomycin, an inhibitor of protein synthesis, its expression does not require prerequisite protein synthesis, indicating that BZLF1 is expressed as an immediate-early gene following primary EBV infection of B lymphocytes.     

Purification, cloning, and expression of an alpha/beta-galactoside alpha-2,3-sialyltransferase from a luminous marine bacterium, Photobacterium phosphoreum

A novel sialyltransferase, alpha/beta-galactoside alpha-2,3-sialyltransferase, was purified from the cell lysate of a luminous marine bacterium, Photobacterium phosphoreum JT-ISH-467, isolated from the Japanese common squid (Todarodes pacificus). The gene encoding the enzyme was cloned from the genomic library of the bacterium using probes derived from the NH(2)-terminal and internal amino acid sequences. An open reading frame of 409 amino acids was identified, and the sequence had 32% identity with that of beta-galactoside alpha-2,6-sialyltrasferase in Photobacterium damselae JT0160. DNA fragments that encoded the full-length protein and a protein that lacked the sequence between the 2nd and 24th residues at the NH(2) terminus were amplified by polymerase chain reactions and cloned into an expression vector. The full-length and truncated proteins were expressed in Escherichia coli, producing active enzymes of 0.25 and 305 milliunits, respectively, per milliliter of the medium in the lysate of E. coli. The truncated enzyme was much more soluble without detergent than the full-length enzyme. The enzyme catalyzed the transfer of N-acetylneuraminic acid from CMP-N-acetylneuraminic acid to disaccharides, such as lactose and N-acetyllactosamine, with low apparent K(m) and to monosaccharides, such as alpha-methyl-galactopyranoside and beta-methyl-galactopyranoside, with much lower apparent K(m). Thus, this sialyltransferase is unique and should be very useful for achieving high productivity in E. coli with a wide substrate range.     

Crystal structures of blasticidin S deaminase (BSD): implications for dynamic properties of catalytic zinc

The set of blasticidin S (BS) and blasticidin S deaminase (BSD) is a widely used selectable marker for gene transfer experiments. BSD is a member of the cytidine deaminase (CDA) family; it is a zinc-dependent enzyme with three cysteines and one water molecule as zinc ligands. The crystal structures of BSD were determined in six states (i.e. native, substrate-bound, product-bound, cacodylate-bound, substrate-bound E56Q mutant, and R90K mutant). In the structures, the zinc position and coordination structures vary. The substrate-bound structure shows a large positional and geometrical shift of zinc with a double-headed electron density of the substrate that seems to be assigned to the amino and hydroxyl groups of the substrate and product, respectively. In this intermediate-like structure, the steric hindrance of the hydroxyl group pushes the zinc into the triangular plane consisting of three cysteines with a positional shift of approximately 0.6 A, and the fifth ligand water approaches the opposite direction of the substrate with a shift of 0.4 A. Accordingly, the zinc coordination is changed from tetrahedral to trigonal bipyramidal, and its coordination distance is extended between zinc and its intermediate. The shift of zinc and the recruited water is also observed in the structure of the inactivated E56Q mutant. This novel observation is different in two-cysteine cytidine deaminase Escherichia coli CDA and might be essential for the reaction mechanism in BSD, since it is useful for the easy release of the product by charge compensation and for the structural change of the substrate.     

Genetic structure of masu salmon (Oncorhynchus masou) populations in Hokkaido, northernmost Japan, inferred from mitochondrial DNA variation

Genetic structure of masu salmon Oncorhynchus masou populations in Hokkaido was examined by analysing mtDNA NADH dehydrogenase subunit 5 gene (561 bp) of 382 individuals collected from 12 rivers, in which there were no records of artificial release. Analysis of molecular variance showed that between groups level and between populations within-group level explained each c. 10% of genetic variance. In neighbour-joining tree, four populations located in southern Hokkaido were clustered into a single group; however, other populations did not form any clear clusters. Fu's F _S, Tajima's D and a mismatch distribution test indicated a sudden expansion of population in the entire population of Hokkaido and the northernmost population of Chiraibetsu, which was genetically close to the southern Hokkaido group. The Sea of Japan and southern rivers, including those of southern Hokkaido, seem to have served as refugia for masu salmon during glacial periods, and their dispersal and straying in interglacial periods affected the genetic structure of masu salmon populations in Hokkaido.     

A role of Toll-IL-1 receptor domain-containing adaptor-inducing IFN-beta in the host response to Pseudomonas aeruginosa lung infection in mice

Toll-IL-1R domain-containing adaptor-inducing IFN-beta (TRIF) is an adaptor molecule that mediates a distinct TLR signaling pathway. Roles of TRIF in the host defense have been primarily associated with virus infections owing to the induction of IFN-alphabeta. In this study, we investigated a role of TRIF in Pseudomonas aeruginosa infection. In vitro, TRIF-deficient mouse alveolar and peritoneal macrophages showed a complete inhibition of RANTES (CCL5) production, severely impaired TNF and KC (CXCL1) production, and reduced NF-kappaB activation in response to P. aeruginosa stimulation. In vivo, TRIF-deficient mice showed a complete inhibition of RANTES production, a severely impaired TNF and KC production, and an efficient MIP-2 and IL-1beta production in the lung following P. aeruginosa infection. This outcome was associated with a delayed recruitment of neutrophils into the airways. These results suggest that TRIF mediates a distinct cytokine/chemokine profile in response to P. aeruginosa infection. P. aeruginosa-induced RANTES production is completely dependent on TRIF pathway in mice. Importantly, TRIF deficiency leads to impaired clearance of P. aeruginosa from the lung during the initial 24-48 h of infection. Thus, TRIF represents a novel mechanism involved in the development of host response to P. aeruginosa infection.