Analysis by molecular cloning of the human class II genes

The HLA class II genes control immune responsiveness to defined antigens; they encode cell surface heterodimers composed of alpha and beta glycopeptides. Recently, cDNA and genomic clones encoding these chains have been isolated, which allows molecular analysis of the class II genes. cDNA clones encoding the alpha chain of the HLA-DR antigen as well as that of another HLA class II antigen have been identified and characterized by nucleotide sequence analysis. These clones have been used as probes to isolate additional class II alpha cDNA clones in cDNA libraries and to identify polymorphisms in genomic DNA. Polymorphic restriction sites have been localized within the HLA-DR alpha gene and used as genetic markers in the analysis of families and of disease (insulin-dependent diabetes mellitus) and control populations. In addition, cDNA clones encoding the DR beta and DC beta chains were used as hybridization probes to identify DNA polymorphism. cDNA clones encoding the DR gamma (Ii) chain have also been identified; unlike the DR alpha and DR beta loci, the DR gamma gene is located on some chromosome other than chromosome 6. The genetic complexity of the human class II alpha and beta loci, as revealed by analysis with cDNA and genomic clones, is greater than that of the murine class II genes. The extent of that complexity will be defined by future work in this area.     

Chromosomal behavior in starfish (Asterina pectinifera) zygotes under the effect of aphidicolin, an inhibitor of DNA polymerase

When calf thymus histones were labeled fluorescently and microinjected into oocytes of the starfish, Asterina pectinifera, the labeled histones visualized chromosomes during maturation division and cleavage. In doing so, we confirmed the previously reported phenomenon that chromosomes became incompetent at the first cleavage in the aphidicolin-treated egg, although cleavage itself took place. Moreover, we found that chromosomes were aligned at the equator of the metaphase spindle of the first cleavage and that they did not separate into two groups at all, but made a lump in the middle of the spindle. Chromosomes finally entered one blastomere, although they did not participate in the following karyokinesis. DNA and microtubules were examined by cytochemistry and immunofluorescence in order to investigate the relation between chromosome movement and the microtubular cytoskeleton. The mitotic apparatus developed and grew in the aphidicolin-treated cells in the same manner as those in normal cells without normal chromatin condensation or chromosome movement during the first cleavage. However, the mitotic apparatus consisted of two asters without the spindle formed at subsequent cleavages. Electron microscopic study revealed that chromosomes did not condense normally and kinetochores were not detected during the first cleavage. These results indicate that the dynamic changes in microtubular structures during mitosis have poor relation with the chromosome behavior such as prophase chromosome condensation and anaphase chromosome movement.     

The adjuvant activity of synthetic N-acetylmuramyl-dipeptide: evidence of initial target cells for the adjuvant activity.

MurNAc-l-Ala-d-isoGln (N-acetylmuramyl-l-alanyl-d-isoglutamine, MDP), a synthetic compound, acts as an adjuvant on the humoral immune response and on the T cell-mediated immune response. In this report, we attempted to directly demonstrate the initial target cells of MDP for its adjuvant activity in vitro by using cell separation procedures.It was demonstrated that MDP enhanced the immune response following direct interaction with antigen-stimulated T and B lymphocytes, but nonstimulated lymphocytes, shortly after triggering by antigen, and that there was no macrophage requirement for MDP to elicite the adjuvant action in the primary anti-SRBC PFC response in vitro. It has also been demonstrated that the adjuvant activity of MDP is due to an enhancing effect which is different from the possible mitogenic activity to spleen cells and MDP replaces neither a function of macrophages, which is substituted by 2-mercaptoethanol nor a helper function of T cells.     

Species richness,abundance and diversity of ichneumonid wasps in Japanese beech forests impacted by sika deer and sawfly herbivory

We investigated the community structure of ichneumonid wasps inhabiting beech forests at six sites in the Tanzawa Mountains of Japan under different magnitudes of impact by sika deer Cervus nippon and beech sawfly Fagineura crenativora on vegetation. Using yellow flight-interception traps, we captured 2,528 ichneumonid wasps representing 367 species in 23 subfamilies. The number of species at each site ranged from 77 to 136 and approximately 80% of these were low-density species (i.e. only one to two individuals captured per site). The number of individuals at each site ranged from 248 to 897, and the percentage of the beech sawfly parasitoids varied widely from 1% to 57%. The numbers of species in parasitoid groups categorized according to their hosts, that is, sawfly (not including the beech sawfly), Lepidoptera, woodborer, fungivore or the others, did not greatly differ among the study sites. Parasitoids attacking herbivorous insects exceeded others in species richness and abundance at all sites. Six sites were classified into four groups in terms of abundance of the host groups when excluding the parasitoids of the beech sawfly, but into only two groups when including these parasitoids. Species diversity and evenness were the highest at the least impacted site even if the beech sawfly parasitoids were excluded from calculation. We suggested some environmental factors, such as groundcover vegetation, abundance of the beech sawfly and structure and age of forest stands, that could have affected the community structure of ichneumonid wasps in the beech forests.     

Cellular Auxin Homeostasis under High Temperature Is Regulated through a SORTING NEXIN1–Dependent Endosomal Trafficking Pathway

High-temperature-mediated adaptation in plant architecture is linked to the increased synthesis of the phytohormone auxin, which alters cellular auxin homeostasis. The auxin gradient, modulated by cellular auxin homeostasis, plays an important role in regulating the developmental fate of plant organs. Although the signaling mechanism that integrates auxin and high temperature is relatively well understood, the cellular auxin homeostasis mechanism under high temperature is largely unknown. Using the Arabidopsis thaliana root as a model, we demonstrate that under high temperature, roots counterbalance the elevated level of intracellular auxin by promoting shootward auxin efflux in a PIN-FORMED2 (PIN2)-dependent manner. Further analyses revealed that high temperature selectively promotes the retrieval of PIN2 from late endosomes and sorts them to the plasma membrane through an endosomal trafficking pathway dependent on SORTING NEXIN1. Our results demonstrate that recycling endosomal pathway plays an important role in facilitating plants adaptation to increased temperature.     

Substrate-mediated proton relay mechanism for the religation reaction in topoisomerase II

The DNA religation reaction of yeast type II topoisomerase (topo II) was investigated to elucidate its metal-dependent general acid/base catalysis. Quantum mechanical/molecular mechanical calculations were performed for the topo II religation reaction, and the proton transfer pathway was examined. We found a substrate-mediated proton transfer of the topo II religation reaction, which involves the 3′ OH nucleophile, the reactive phosphate, water, Arg781, and Tyr782. Metal A stabilizes the transition states, which is consistent with a two-metal mechanism in topo II. This pathway may be required for the cleavage/religation reaction of topo IA and II and will provide a general explanation for the catalytic mechanism in the topo IA and II.     

Synthesis and Physiological Activity of Methyl 6′6′-Didemethyl Abscisate and New Chiral Analogs

Methyl 6′, 6′-didemethyl abscisate (5) was synthesized and assayed to elucidate the physiological activity of methyl substituents on the cyclohexene ring of abscisic acid (ABA). During this study two new chiral stereoisomeric analogs 6 and 7 were synthesized from l-and d-carvone. The rice seedling assay and germination assay of garden radish showed that 6′-methyl groups of ABA were not important in biological activity and that 5′-isopropenyl analogs 6 and 7 were inactive.     

Syntheses and Biological Activities of 1-O-Glucopyranosyl Fatty Acid Esters

1-O-Palmitoyl-d-glucopyranose was prepared by the selective 1-O-acylation of 4,6-O-benzylideneglucose followed by hydrogenolysis of the protecting group. 1-O-Oleoyl-d-glucopyranose was synthesized from the corresponding benzylidene derivative by selective hydrolysis in acetic acid. This procedure constitutes a useful method for the synthesis of 1-O-acyl-d-glucopyranoses containing unsaturated carboxylic acids. However, 4,6-O-benzylidene-l-O-linolenoyl-d-glucopyranose was converted to 3-O-linolenoyl-d-glucopyranose by the acidic hydrolysis due to acyl migration.

Synthesized glucosyl esters were inactive in the bean second-internode bioassay. However, it was found that 3-O-linolenoyl-d-glucopyranose had a promoting activity on germination of pollen and growth of pollen tube.     

Preparation of Optically Pure cis-2,4-Dimethyl-l-cyclohexanones,the Key Intermediates in Cycloheximide Synthesis,Using Microbial Resolution

Asymmetric hydrolysis of acetate (10) of (±)-t-2,t-4-dimethyl-r-l-cyclohexanol with Bacillus subtilis var. niger gave (?)-(lS,2S,4S)-2,4-dimethyl-l-cyclohexanol (6a) and (+)-(1R,2R,4R)-acetate (10b) with high optical purities. Optically pure (?) and (+)-alcohols (6a and 6b) were prepared via corresponding 3,5-dinitrobenzoates. Oxidation of alcohols (6a and 6b) with chromic acid gave optically pure (?)-(2S,4S) and (+)-(2R,4R)-2,4-dimethyl-l-cyclohexanones (2a and 2b), respectively.