Thymus, innate immunity and autoimmune arthritis: interplay of gene and environment

A hypomorphic mutation of the gene encoding zeta-associated protein-70 (ZAP-70), a signaling molecule in T cells, produces autoimmune arthritis in mice under a microbially conventional condition but not in a clean environment. The genetic anomaly alters thymic selection of self-reactive T cells as well as natural regulatory T cells and their respective functions. Highly self-reactive polyclonal T cells, including arthritogenic ones, thus produced by the thymus strongly recognize self-antigens presented by antigen-presenting cells, stimulate them to up-regulate co-stimulatory molecules and secrete cytokines that drive na?ve self-reactive T cells to differentiate into autoimmune effector Th17 cells. Administration of microbial products and activation of complement can facilitate the differentiation, evoking clinically overt arthritis in a microbially clean environment. Furthermore, mutation-dependent graded attenuation of T cell receptor signaling alters disease phenotypes and the dependency of disease occurrence on the environment. These findings provide a model of how genetic and environmental factors, in association, cause autoimmune diseases such as rheumatoid arthritis.     

Differentiation in wild-type allele of the sd1 locus concerning culm length between indica and japonica subspecies of Oryza sativa (rice)

A dwarfing gene (allele) sd1-d has been intensively utilized to develop short-culm indica varieties in southeast Asia up to now. Before the first sd1-d-carrying variety IR8 was released, rice researchers had recognized the general tendency that culm length is higher in indica varieties than in temperate-japonica ones. Inter-subspecific difference of the tall (wild-type) allele SD1 at the sd1 locus was examined on the common genetic background, using five isogenic lines developed by substituting sd1-d of the recurrent parent IR36 by SD1s of two indica varieties, two temperate-japonica varieties and one tropical-japonica variety. The two indica -donor isogenic lines had longer culms than the three japonica-donor isogenic lines consistently in two different environmental conditions. Moreover, nonsynonymous single-nucleotide polymorphism between the two subspecies was detected at two sites in Exon 1 and Exon 3 of the sd1 locus. It is demonstrated that the inter-subspecific differentiation of SD1 contributes height difference between indica and japonica. The indica-originating and japonica-originating alleles at the sd1 locus were designated as SD1-in(t) and SD1-ja(t), respectively.     

Axenic culture of <Emphasis Type="Italic">Brachionus plicatilis</Emphasis> using antibiotics

The rotifer Brachionus plicatilis culture is composed of complex microcosms including bacteria, protozoans, algae, and fungi. Previous studies reported methods to establish axenic rotifer cultures, but further refinement of these techniques is needed, for molecular biological research which requires pure culture to isolate nucleic acids from rotifers only. In order to render rotifer culture axenic, we tested five antibiotics: ampicillin (Amp), chloramphenicol (Cp), kanamycin (Km), nalidixic acid (Na), and streptomycin (Sm) at 30–100 μg/ml. Except for Cp, which reduces rotifer reproduction, all other antibiotics at the tested concentrations did not affect rotifer reproduction or show any toxic effects. A rotifer disinfection method was finally established by treating the resting eggs with 0.25% (w/v) sodium hypochlorite (NaOCl) for 3 min, washing with sterilized sea water, and then exposing the neonates to an Amp, Km, Na, and Sm mixture. Using four nutrient media, we confirmed that this protocol renders the rotifer culture bacterial and fungus free. The axenic rotifer culture generated here is useful not only for genetic analysis of Brachionus plicatilis, but for studying the rotifer life cycle without bacterial influence.     

Sequence analysis of genomic DNA (680 Mb) by GS-FLX-Titanium sequencer in the monogonont rotifer, Brachionus ibericus

The monogonont rotifer, Brachionus ibericus (S type), is considered to be a promising model species for developmental biology, evolution, and environmental genomics. In an attempt to accelerate the molecular understanding of B. ibericus, we sequenced 680.5 Mb of genomic DNA using the genome sequencer GS-FLX-Titanium. We obtained 2,062,621 reads (average read length 329.9 bp) and 145,418 contigs (total contigs length 125.7 Mb) after excluding small reads (less than 200 bp) from the assembly, and finally obtained 10,133 unigenes (E value ?? 9.00E?04) after non-redundant (NR) BLAST search. In this article, we summarize the genomic DNA sequences of B. ibericus and discuss their potential use in the study of reproductive biology, endocrinology, environmental genomics, and ecotoxicological studies, and for providing insight into the genetic basis of mechanisms such as egg formation, antioxidant stress defense, and xenobiotic metabolism.     

Synthesis of blockwise alkylated (1→4) linked trisaccharides as surfactants: influence of configuration of anomeric position on their surface activities

New carbohydrate-based surfactants consisting of hydrophilic cellobiosyl and hydrophobic glucosyl residues, methyl β-d-glucopyranosyl-(1→4)-α-d-glucopyranosyl-(1→4)-2,3,6-tri-O-methyl-α-d-glucopyranoside 1 (GβGαMα, G: glucopyranosyl residue, α and β: α-(1→4)- and β-(1→4) glycosidic bonds, M: methyl group), 2 (G(β)G(β)M(α)), 3 (G(β)G(α)M(β)), 4 (G(β)G(β)M(β)), 5 (G(β)G(α)E(α), E: ethyl group), 6 (G(β)G(β)E(α)), 7 (G(β)G(α)E(β)), 8 (G(β)G(β)E(β)) and eight α-and β-glycoside mixtures (a mixture of 1 and 2: 1/2=62/38 (9), 32/68 (10); a mixture of 3 and 4: 3/4=69/31 (11), 32/68 (12); a mixture of 5 and 6: 5/6=62/38 (13), 33/67 (14); a mixture of 7 and 8: 7/8=59/41 (15), 29/71 (16)) were synthesized via combined methods consisting of acid-catalyzed alcoholysis of cellulose ethers and glycosylation of phenyl thio-cellobioside derivatives. Their surface activities in aqueous solution depended on their chemical structures: α- or β-(1→4) linkage between hydrophilic cellobiosyl and hydrophobic glucosyl blocks, methyl or ethyl groups of hydrophobic glucosyl block, and α- or β-linked ether group at the C-1 of hydrophobic glucosyl block. The mixing effect of α- and β-glycosides on surface activities was also investigated. As a result, ethyl β-d-glucopyranosyl-(1→4)-α-d-glucopyranosyl-(1→4)-2,3,6-tri-O-ethyl-β-d-glucopyranoside 7 (G(β)G(α)E(β)) had the highest surface activity, and its critical micellar concentration (CMC) and γ(CMC) (surface tension at CMC) values of compound 7 were 0.5mM (ca. 0.03wt%) and 34.5mN/m, respectively. The surface tensions of α- and β-glycoside mixtures except for compounds 9 and 10 were almost equal to those of pure compounds. The syntheses of the mixtures of α- and β-glycosides without purification process are easier than those of pure compounds. Thus, the mixtures should be more practical compounds for industrial use as a surfactant.     

Monocarboxylate transporter-1 is required for cell death in mouse chondrocytic ATDC5 cells exposed to interleukin-1beta via late phase activation of nuclear factor kappaB and expression of phagocyte-type NADPH oxidase

Interleukin-1β (IL-1β) induces cell death in chondrocytes in a nitric oxide (NO)- and reactive oxygen species (ROS)-dependent manner. In this study, increased production of lactate was observed in IL-1β-treated mouse chondrocytic ATDC5 cells prior to the onset of their death. IL-1β-induced cell death in ATDC5 cells was suppressed by introducing an siRNA for monocarboxylate transporter-1 (MCT-1), a lactate transporter distributed in plasma and mitochondrial inner membranes. Mct-1 knockdown also prevented IL-1β-induced expression of phagocyte-type NADPH oxidase (NOX-2), an enzyme specialized for production of ROS, whereas it did not have an effect on inducible NO synthase. Suppression of IL-1β-induced cell death by Nox-2 siRNA indicated that NOX-2 is involved in cell death. Phosphorylation and degradation of inhibitor of κBα (IκBα) from 5 to 20 min after the addition of IL-1β was not affected by Mct-1 siRNA. In addition, IκBα was slightly decreased after 12 h of incubation with IL-1β, and the decrease was prominent after 36 h, whereas activation of p65/RelA was observed from 12 to 48 h after exposure to IL-1β. These changes were not seen in Mct-1-silenced cells. Forced expression of IκBα super repressor as well as treatment with the IκB kinase inhibitor BAY 11-7082 suppressed NOX-2 expression. Furthermore, Mct-1 siRNA lowered the level of ROS generated after 15-h exposure to IL-1β, whereas a ROS scavenger, N-acetylcysteine, suppressed both late phase degradation of IκBα and Nox-2 expression. These results suggest that MCT-1 contributes to NOX-2 expression via late phase activation of NF-κB in a ROS-dependent manner in ATDC5 cells exposed to IL-1β.     

Role of AtPolζ, AtRev1, and AtPolη in UV light-induced mutagenesis in Arabidopsis

Translesion synthesis (TLS) is a DNA damage tolerance mechanism in which DNA lesions are bypassed by specific polymerases. To investigate the role of TLS activities in ultraviolet light-induced somatic mutations, we analyzed Arabidopsis (Arabidopsis thaliana) disruptants of AtREV3, AtREV1, and/or AtPOLH genes that encode TLS-type polymerases. The mutation frequency in rev3-1 or rev1-1 mutants decreased compared with that in the wild type, suggesting that AtPolζ and AtRev1 perform mutagenic bypass events, whereas the mutation frequency in the polh-1 mutant increased, suggesting that AtPolη performs nonmutagenic bypass events with respect to ultraviolet light-induced lesions. The rev3-1 rev1-1 double mutant showed almost the same mutation frequency as the rev1-1 single mutant. The increased mutation frequency found in polh-1 was completely suppressed in the rev3-1 polh-1 double mutant, indicating that AtPolζ is responsible for the increased mutations found in polh-1. In summary, these results suggest that AtPolζ and AtRev1 are involved in the same (error-prone) TLS pathway that is independent from the other (error-free) TLS pathway mediated by AtPolη.