Antitumour activities of levans produced by Zymomonas mobilis strains

Levans produced by four Zymomonas mobilis strains showed antitumour activity against sarcoma 180 and Ehrlich carcinoma in Swiss albino mice. Levans from two strains (ZAP and CP4) had the highest effects. NMR analysis showed that the polymers were composed only of fructose units. The results suggested that the antineoplasic effect is associated to the polysaccharide molecular weight and that a particular molecular weight range may be responsible for this effect.     

Catabolism of tissue-type plasminogen activator by the human hepatoma cell line Hep G2. Modulation by plasminogen activator inhibitor type 1

Catalytic activity of tissue-type plasminogen activator (t-PA) in plasma is regulated in part by formation of complexes with specific inhibitors as well as by hepatic clearance. Potential interaction of these two regulatory mechanisms was examined in the human hepatoma cell line Hep G2. These cells secrete plasminogen activator inhibitor type-1 (PAI-1) and initiate catabolism of exogenous t-PA by receptor-mediated endocytosis. Specific binding of 125I-t-PA to cells at 4 degrees C results in dose-dependent formation of a 95-kDa species recognized by monospecific anti-PAI-1 and anti-t-PA antibodies and stable in the presence of low (0.2%) concentrations of sodium dodecyl sulfate (SDS). Specific binding of 125I-t-PA and formation of the 95-kDa SDS-stable species are inhibited in a concentration-dependent manner following preincubation of cells with anti-PAI-1 antibodies. High and low molecular weight forms of urokinase plasminogen activator (u-PA) capable of forming specific complexes with PAI-1 complete for 125I-t-PA binding sites. However, the proenzyme form of u-PA (scu-PA), incapable of forming complexes with PAI-1, does not compete for 125I-t-PA binding sites. The role of the serine protease active site of t-PA in mediating both interaction with PAI-1 and specific binding was examined using 125I-t-PA that had been functionally inactivated with D-phenylalanyl-L-propyl-L-arginyl-chloromethyl ketone (PPACK). 125I-t-PA-PPACK, despite a 6-fold lower affinity than active 125I-t-PA, exhibited specific binding to cells without detectable formation of SDS-stable complexes with PAI-1. Both surface-bound 125I-t-PA and 125I-t-PA-PPACK are internalized and degraded by cells at 37 degrees C. 125I-t-PA is internalized as a stable complex with PAI-1, whereas 125I-t-PA-PPACK is internalized with similar kinetics but without the presence of an SDS-stable complex. Thus, PAI-1 appears capable of modulating t-PA catabolism in the human hepatocyte.     

Biomass production in a nitrogen-fertilized,tallgrass prairie ecosystem exposed to ambient and elevated levels of CO2

Increased biomass production in terrestrial ecosystems with elevated atmospheric CO₂ may be constrained by nutrient limitations as a result of increased requirement or reduced availability caused by reduced turnover rates of nutrients. To determine the short-term impact of nitrogen (N) fertilization on plant biomass production under elevated CO₂, we compared the response of N-fertilized tallgrass prairie at ambient and twice-ambient CO₂ levels over a 2-year period. Native tallgrass prairie plots (4.5 m diameter) were exposed continuously (24 h) to ambient and twice-ambient CO₂ from 1 April to 26 October. We compared our results to an unfertilized companion experiment on the same research site. Above- and belowground biomass production and leaf area of fertilized plots were greater with elevated than ambient CO₂ in both years. The increase in biomass at high CO₂ occurred mainly aboveground in 1991, a dry year, and belowground in 1990, a wet year. Nitrogen concentration was lower in plants exposed to elevated CO₂, but total standing crop N was greater at high CO₂. Increased root biomass under elevated CO₂ apparently increased N uptake. The biomass production response to elevated CO₂ was much greater on N-fertilized than unfertilized prairie, particularly in the dry year. We conclude that biomass production response to elevated CO₂ was suppressed by N limitation in years with below-normal precipitation. Reduced N concentration in above- and belowground biomass could slow microbial degradation of soil organic matter and surface litter, thereby exacerbating N limitation in the long term.     

Variation in the ribosomal internal transcribed spacers (ITS1 and ITS2) among eight taxa of the Mimulus guttatus species complex

The internal transcribed spacer region (ITS1 and ITS2) of the 18S-25S nuclear ribosomal DNA sequence and the intervening 5.8S region were sequenced from three individuals in each of eight taxa of the Mimulus guttatus species complex. Three discrete variants, or "types," of ITS sequences were found, among which 30%-40% of sites differed, compared with 1%-2% within types. Dot plots indicate that these types were not related by conspicuous rearrangements or inversions. More than one ITS type was often found in the same taxon, and two of three ITS types span species boundaries, indicating their presence prior to speciation. These ITS sequences showed essentially no positional homology with the nearest sequenced relative, tomato. In contrast, the 5.8S region was relatively unvaried, with 8 of 162 sites varied in the sample among all eight taxa. The phylogeny inferred by the most common ITS sequence type, rooted by the two other ITS types, agreed with isozymes in showing the distinctness of M. nudatus, M. laciniatus, and M. tilingii from the other five taxa.      

Responses in stomatal conductance to elevated CO2 in 12 grassland species that differ in growth form

Responses in stomatal conductance (g _st ) and leaf xylem pressure potential ( _leaf ) to elevated CO₂ (2x ambient) were compared among 12 tallgrass prairie species that differed in growth form and growth rate. Open-top chambers (OTCs, 4.5 m diameter, 4.0 m in height) were used to expose plants to ambient and elevated CO₂ concentrations from April through November in undisturbed tallgrass prairie in NE Kansas (USA). In June and August, _leaf was usually higher in all species at elevated CO₂ and was lowest in adjacent field plots (without OTCs). During June, when water availability was high, elevated CO₂ resulted in decreased g _st in 10 of the 12 species measured. Greatest decreases in g _st (ca. 50%) occurred in growth forms with the highest potential growth rates (C₃ and C₄ grasses, and C₃ ruderals). In contrast, no significant decrease in g _st was measured in the two C₃ shrubs. During a dry period in September, reductions in g _st at elevated CO₂ were measured in only two species (a C₃ ruderal and a C₄ grass) whereas increased g _st at elevated CO₂ was measured in the shrubs and a C₃ forb. These increases in g _st were attributed to enhanced _leaf in the elevated CO₂ plants resulting from increased soil water availability and/or greater root biomass. During a wet period in September, only reductions in g _st were measured in response to elevated CO₂. Thus, there was significant interspecific variability in stomatal responses to CO₂ that may be related to growth form or growth rate and plant water relations. The effect of growth in the OTCs, relative to field plants, was usually positive for g _st and was greatest (>30%) when water availability was low, but only 6–12% when _leaf was high.The results of this study confirm the importance of considering interactions between indirect effects of high CO₂ of plant water relations and direct effects of elevated CO₂ on g _st , particularly in ecosystems such as grasslands where water availability often limits productivity. A product of this interaction is that the potential exists for either positive or negative responses in g _st to be measured at elevated levels of CO₂.     

Variation in heat shock proteins within tropical and desert species of poeciliid fishes

The 70-kilodalton heat shock protein (hsp70) family of molecular chaperones, which contains both stress-inducible and normally abundant constitutive members, is highly conserved across distantly related taxa. Analysis of this protein family in individuals from an outbred population of tropical topminnows, Poeciliopsis gracilis, showed that while constitutive hsp70 family members showed no variation in protein isoforms, inducibly synthesized hsp70 was polymorphic. Several species of Poeciliopsis adapted to desert environments exhibited lower levels of inducible hsp70 polymorphism than the tropical species, but constitutive forms were identical to those in P. gracilis, as they were in the confamilial species Gambusia affinis. These differences suggest that inducible and constitutive members of this family are under different evolutionary constraints and may indicate differences in their function within the cell. Also, northern desert species of Poeciliopsis synthesize a subset of the inducible hsp70 isoforms seen in tropical species. This distribution supports the theory that ancestral tropical fish migrated northward and colonized desert streams; the subsequent decrease in variation of inducible hsp70 may have been due to genetic drift or a consequence of adaptation to the desert environment. Higher levels of variability were found when the 30- kilodalton heat shock protein (hsp30) family was analyzed within different strains of two desert species of Poeciliopsis and also in wild-caught individuals of Gambusia affinis. In both cases the distribution of hsp30 isoform diversity was similar to that seen previously with allozyme polymorphisms.      

Binding of plasminogen activator inhibitor type-1 to extracellular matrix of Hep G2 cells. Evidence that the binding protein is vitronectin.

Catabolism of plasminogen activators by Hep G2 cells is mediated by a specific receptor which recognizes complexes of these serine proteases with their physiological inhibitor, plasminogen activator inhibitor type-1 (PAI-1). This catabolic process is initiated by interaction of exogenous plasminogen activators with bioactive PAI-1, which is secreted and localizes in an active form to the extracellular matrix (ECM) of Hep G2 cells. We now report that vitronectin (VN) mediates the specific binding of PAI-1 to the ECM of these cells. Purified bovine or human VN competes for specific binding of PAI-1 to Hep G2 ECM, and ligand blotting reveals specific binding of PAI-1 to ECM-associated VN. Hep G2 cells secrete both VN and PAI-1, and pulse-chase studies strongly suggest that these proteins associate only following secretion. Although Hep G2 cell-derived VN does not significantly bind to ECM in vitro, 30-40% of endogenous PAI-1 binds to the ECM, even in the presence of human serum, suggesting that ECM-associated VN is entirely derived from bovine serum. PAI-1 was localized by indirect immunofluorescence to ECM beneath cells and at cell margins, whereas VN exhibited a uniform distribution throughout the growth substratum. VN associated with the ECM may confer retention and bioactivity to PAI-1, potentially facilitating both pericellular regulation of plasmin generation and the rapid hepatic clearance of plasminogen activators.     

Elevated CO22 and Leaf Longevity in the C44 Grassland-Dominant Andropogon gerardii

In central U.S. grasslands, plant and ecosystem responses to elevated CO2 are most pronounced when water availability is limited. In a northeast Kansas grassland, responses to elevated CO2 in leaf area, number, development, and longevity were quantified for the tallgrass prairie dominant, Andropogon gerardii. Plants were grown in open-top chambers (OTCs) modified to limit water availability and to maximize responses to elevated CO2. In OTCs with elevated (x2 ambient) levels of CO2, aboveground biomass production and leaf water potentials were increased significantly compared with those of plants in OTCs with ambient CO2. There were no differences in leaf area or leaf number per tiller in A. gerardii in elevated compared with ambient OTCs. However, leaf area in adjacent unchambered plots with greater water availability was significantly higher than in the OTCs. The time required for developing leaves to achieve maximum leaf area was reduced by 29%, and the period of time until leaves senesced was increased by 20% for plants exposed to elevated compared with ambient CO2. Thus, leaves of this C4 grass species expanded more rapidly (6 d) and remained green longer (9 d) when exposed to elevated CO2. Such CO2-mediated increases in leaf longevity in the dominant species may allow this grassland to respond more opportunistically to temporally variable rainfall patterns in high-CO2 environments. These responses should be included in leaf-based simulation models that attempt to mechanistically link physiological alterations to predicted canopy responses to increased CO2.     

柠檬酸合酶的分子生物学研究进展

柠檬酸合酶（citrate synthase,CS）是细胞内多种重要代谢途径的关键酶。CS可催化草酰乙酸和乙酰辅酶A之间的缩合反应生成柠檬酸和辅酶A。通常革兰氏阳性细菌、古菌以及真核细胞的CS为同源二聚体,而革兰氏阴性细菌的CS为同源六聚体。根据其在细胞内的定位不同,CS可分为线粒体CS、乙醛酸循环体CS、过氧化物酶体CS。这些同工酶在能量代谢、植物脂肪的代谢、脂肪酸的氧化及细胞解毒过程中起着重要作用。不同来源的CS空间结构、催化机制和动力学性质十分相似。针对其生化特性、空间结构特点、催化机制以及分子进化等研究进展进行综述。     

Effects of long-term fixation on histological quality of undecalcified murine bones embedded in methylmethacrylate

While long-term fixation and storage of specimens is common and useful for many research projects, it is particularly important for space flight investigations where samples may not be returned to Earth for several months (International Space Station) or years (manned mission to Mars). We examined two critical challenges of space flight experimentation: the effect of long-term fixation on the quality of mouse bone preservation and the preservation of antigens and enzymes for both histochemical and immunohistochemical analyses, and how the animal/sample processing affects the preservation. We show that long-term fixation minimally affects standard histological staining, but that enzyme histochemistry and immunolabeling are greatly compromised. Further, we demonstrate that whole animal preservation is not as suitable as whole leg or stripped leg preservation for long-term fixation and all histological analyses. Overall, we recommend whole leg processing for long-term storage of bone specimens in fixative prior to embedding in plastic for histological examination.