Adrenomedullin binding protein-1 modulates vascular responsiveness to adrenomedullin in late sepsis

Adrenomedullin (AM), a potent vasodilatory peptide, plays an important role in initiating the hyperdynamic response during the early stage of sepsis. Moreover, the reduced vascular responsiveness to AM appears to be responsible for the transition from the early, hyperdynamic to the late, hypodynamic phase of sepsis. Although the novel specific AM binding protein-1 (AMBP-1) enhances AM-mediated action in a cultured cell line, it remains to be determined whether AMBP-1 plays any role in modulating vascular responsiveness to AM during sepsis. To study this, adult male rats were subjected to sepsis by cecal ligation and puncture (CLP). The thoracic aorta was harvested for determination of AM-induced vascular relaxation. Aortic levels of AMBP-1 were determined by Western blot analysis, and AM receptor gene expression in the aortic tissue was assessed by RT-PCR. The results indicate that AMBP-1 significantly enhanced AM-induced vascular relaxation in aortic rings from sham-operated animals. Although vascular responsiveness to AM decreased at 20 h after CLP (i.e., the late, hypodynamic stage of sepsis), addition of AMBP-1 in vitro restored the vascular relaxation induced by AM. Moreover, the aortic level of AMBP-1 decreased significantly at 20 h after CLP. In contrast, AM receptor gene expression was not altered under such conditions. These results, taken together, suggest that AMBP-1 plays an important role in modulating vascular responsiveness to AM, and the reduced AMBP-1 appears to be responsible for the vascular AM hyporesponsiveness observed during the hypodynamic phase of sepsis.     

Oncogenic activation of the human trk proto-oncogene by recombination with the ribosomal large subunit protein L7a.

The trk-2h oncogene, isolated from the human breast carcinoma cell line MDA-MB 231 by genomic DNA-transfection into NIH3T3 cells, consists of the trk proto-oncogene receptor kinase domain fused to a N-terminal 41 amino acid activating sequence (Kozma, S.C., Redmond, S.M.S., Xiao-Chang, F., Saurer, S.M., Groner, B. and Hynes, N.E. (1988) EMBO J., 7, 147-154). Antibodies raised against a bacterially produced beta gal-trk receptor kinase fusion protein recognized a 44 kd phosphoprotein phosphorylated on serine, threonine and tyrosine in extracts of trk-2h transformed NIH3T3 cells. In vitro, in the presence of Mn2+/gamma ATP, this protein became phosphorylated extensively on tyrosine. Cells transformed by trk-2h did not, however, show an elevation in total phosphotyrosine. We have cloned and sequenced the cDNA encoding the amino terminal activating sequences of trk-2h (Kozma et al., 1988). The encoded protein has a high basic amino acid content and the gene is expressed as an abundant 1.2 kb mRNA in human, rat and mouse cells. Antipeptide antibodies raised against a C-terminal peptide recognized specifically a 30 kd protein on Western blots of human, rat and mouse cell extracts. Immunofluorescence revealed, in addition to granular cytoplasmic fluorescence, intense nucleolar staining. The high basic amino acid content and nucleolar staining prompted us to investigate whether the 30 kd protein could be a ribosomal protein. Western immunoblotting analysis of 2D-electrophoretically resolved ribosomal proteins indicated that the 30 kd protein is the ribosomal large subunit protein L7a.(ABSTRACT TRUNCATED AT 250 WORDS)     

SWATH‐based quantitative proteomics reveals the mechanism of enhanced Bombyx mori nucleopolyhedrovirus‐resistance in silkworm reared on UV‐B treated mulberry leaves

Bombyx mori nucleopolyhedrovirus (BmNPV) is one of the most acute infectious diseases in silkworm, which has led to great economic loss in sericulture. Previous study showed that the content of secondary metabolites in mulberry leaves, particularly for moracin N, was increased after UV‐B irradiation. In this study, the BmNPV resistance of silkworms reared on UV‐B treated and moracin N spread mulberry leaves was improved. To uncover the mechanism of enhanced BmNPV resistance, silkworm midguts from UV‐B treated mulberry leaves (BUM) and moracin N (BNM) groups were analyzed by SWATH‐based proteomic technique. Of note, the abundance of ribosomal proteins in BUM and BNM groups was significantly changed to maintain the synthesis of total protein levels and cell survival. While, cytochrome c oxidase subunit II, calcium ATPase and programmed cell death 4 involved in apoptotic process were up‐regulated in BNM group. Expressions of lipase‐1, serine protease precursor, Rab1 protein, and histone genes were increased significantly in BNM group. These results suggest that moracin N might be the main active component in UV‐B treated mulberry leaves which could improve the BmNPV‐resistance of silkworm through promoting apoptotic cell death, enhancing the organism immunity, and regulating the intercellular environment of cells in silkworm. It also presents an innovative process to reduce the mortality rate of silkworms infected with BmNPV.     

The Cenomanian–Turonian boundary in the northwestern part of the Adriatic Carbonate Platform (Ćićarija Mtn., Istria,Croatia): characteristics and implications

The Cenomanian–Turonian boundary (CTB) in the ?i?arija Mountain region (northern Istria, Croatia) is characterized by calcisphere limestone successions with a firmground and glauconite horizon, bioturbated intervals, tempestites, and slumped structures as well as microbially laminated and organic-rich interbeds deposited in the northwestern part of the intra-Tethyan Adriatic Carbonate Platform (AdCP). Compilation of the results from three studied sections (Vodice–Jelovica, Martinjak and Planik) of litho-, bio-, and microfacies analyses, X-ray diffraction, SEM, EDS, and stable isotope analyses allowed reconstruction of marine paleoenvironmental conditions during this time period. Shallow-marine carbonate deposits of the Milna Formation underlie a drowned-platform succession of the Sveti (Sv.) Duh Formation. The contact between these two formations is sharp and commonly marked by slumped deposits. The Sv. Duh Formation consists of about 100 m of calcisphere wackestone enriched in organic matter. The results of preliminary δ¹³C and δ¹⁸O stable isotope analyses indicate the influence of the global Oceanic Anoxic Event (OAE2) on the deposition of this carbonate succession. Anoxic and hypoxic conditions in the water column lead to major changes in the shallow-marine carbonate system of the AdCP. Numerous benthic foraminifera declined during that time, but planktonic foraminifera and calcareous dinoflagellates diversified and expanded greatly. The results of this research provide new insights into the character of the CTB interval in this part of the Tethyan realm. Local and regional synsedimentary tectonics combined with global upper Cretaceous sea-level dynamics allows the correlation of the investigated deeper-marine lithostratigraphic units with OAE2.     

Stochastic context-free grammars for tRNA modeling.

Stochastic context-free grammars (SCFGs) are applied to the problems of folding, aligning and modeling families of tRNA sequences. SCFGs capture the sequences' common primary and secondary structure and generalize the hidden Markov models (HMMs) used in related work on protein and DNA. Results show that after having been trained on as few as 20 tRNA sequences from only two tRNA subfamilies (mitochondrial and cytoplasmic), the model can discern general tRNA from similar-length RNA sequences of other kinds, can find secondary structure of new tRNA sequences, and can produce multiple alignments of large sets of tRNA sequences. Our results suggest potential improvements in the alignments of the D- and T-domains in some mitochondrial tRNAs that cannot be fit into the canonical secondary structure.     

Influence of amino acids on nitrogen fixation ability and growth of Azospirillum spp.

The utilization of amino acids for growth and their effects on nitrogen fixation differ greatly among the several strains of each species of Azospirillum spp. that were examined. A. brasiliense grew poorly or not at all on glutamate, aspartate, serine, or histidine as the sole nitrogen and carbon sources. Nitrogen fixation by most A. brasiliense strains was inhibited only slightly even by 10 mM concentrations of these amino acids. In contrast, A. lipoferum and A. amazonense grew very well on glutamate, aspartate, serine, or histidine as the sole nitrogen and carbon sources; nitrogen fixation, which was measured in the presence of malate or sucrose, was severely inhibited by these amino acids. It was concluded that growth on histidine as the sole source of nitrogen, carbon, and energy may be used for the taxonomic characterization of Azospirillum spp. and for the selective isolation of A. lipoferum. The different utilization of various amino acids by Azospirillum spp. may be important for their establishment in the rhizosphere and for their associative nitrogen fixation with plants. The physiological basis for the different utilization of glutamate by Azospirillum spp. was investigated further. A. brasiliense and A. lipoferum exhibited a high affinity for glutamate uptake (Km values for uptake were 8 and 40 microM, respectively); the Vmax was 6 times higher in A. lipoferum than in A. brasiliense. At high substrate concentrations (10 mM), the nonsaturable component of glutamate uptake was most active in A. lipoferum and A. amazonense.(ABSTRACT TRUNCATED AT 250 WORDS)     

Trypsin-sensitive neutralization site on VP1 of Theiler''s murine encephalomyelitis viruses.

We generated Theiler's murine encephalomyelitis virus mutants resistant to several neutralizing monoclonal antibodies (MAbs) having their epitopes near a trypsin cleavage site of VP1. Neutralization and Western blot (immunoblot) studies suggest that two of the MAbs have identical epitopes that partly overlap the epitope of a third MAb. Sequencing of RNA of the mutants localized the epitopes to a site near the carboxyl end of VP1. The limited diversity of nucleotide changes seen in the mutants and the immunodominance of the site suggest that the carboxyl end of VP1 may have an important function.     

Soil Seedbank in a Dipterocarp Rain Forest in Xishuangbanna,Southwest China1

Dipterocarp rain forest reaches its northern latitudinal limit in Xishuangbanna, Southwest China. We studied the soil seedbank of dipterocarp rain forest in Xishuangbanna during the dry and wet seasons. Results showed that there were large seed accumulations in both the dry (mean ± SD; 3925 ± 2533 seeds/m²) and wet seasons (5415 ± 3232 seeds/m²). One hundred and sixteen species of seed plants were identified from germination, 66 percent of which were woody species. Weed or pioneer species dominated the seedbank. The soil seedbank in Xishuangbanna had similar species composition as compared to those in tropical Asia, but higher seed storage reflects the intense disturbance and forest fragmentation in this area.     

Analysis of cell-growth-phase-related variations in hyaluronate synthase activity of isolated plasma-membrane fractions of cultured human skin fibroblasts.

Hyaluronate synthase activity is localized exclusively in plasma-membrane fractions of cultured human skin fibroblasts. The enzyme activity of plasma membranes prepared from exponential-growth-phase cells was about 6.5 times that of stationary-growth-phase cells. Hyaluronate synthase from exponential-growth-phase cells exhibited lower Km and higher Vmax. values for both UDP-N-acetylglucosamine and UDP-glucuronic acid and higher rate of elongation of hyaluronate chains compared with the enzyme from stationary-growth-phase cells. Hyaluronate synthase exhibited an extremely short half-life, 2.2 h and 3.8 h respectively when cells were treated with cycloheximide and actinomycin D. The cell-growth-phase-dependent variations in hyaluronate synthase activity appear to be due to its high turnover rate as well as due to some post-translational modification of the enzyme protein as cells progress from early exponential to stationary growth phase. The isolated plasma membranes contained a protein (Mr approx. 450,000) that was selectively autophosphorylated from [gamma-32P]ATP in vitro in the presence of hyaluronate precursors in the reaction mixture and that also exhibited some hyaluronate-synthesis-related properties. The 32P-labelled protein isolated from plasma membranes of exponentially growing cells expressed an efficient UDP-[14C]glucuronic acid- and UDP-N-acetyl[3H]glucosamine-binding activity and was able to synthesize oligosaccharides (Mr 5000) of [14C]glucuronic acid and N-acetyl[3H]glucosamine residues. The corresponding protein of stationary-growth-phase cells, which expressed much higher nucleotide-sugar-precursor-binding activity, appeared to have lost its oligosaccharide-synthesizing activity.