Hybridization properties of support-bound oligonucleotides: the effect of the site of immobilization on the stability and selectivity of duplex formation

Four 12-mer oligodeoxyribonucleotide sequences were immobilized to uniformly sized (50 microm) polymer particles through C5-tethered thymine and N(4)-tethered cytosine bases at four different sites in each sequence. The effect of the site of immobilization on the efficiency and selectivity of hybridization of the particle-bound probes was quantified by a sandwich-type assay based on a time-resolved fluorometric measurement of an oligonucleotide probe labeled with a photoluminescent europium(III) chelate directly from the surface of a single particle. Immobilization through a base in the central part of the sequence was observed to destablize the duplex more markedly than tethering through a terminal base. The effect of a one-base mismatch on the duplex stability increased with the increasing distance from the site of immobilization.     

Structure-activity relationships for a series of thiobenzamide influenza fusion inhibitors derived from 1,3,3-trimethyl-5-hydroxy-cyclohexylmethylamine

The anti-influenza activity of a series of thiobenzamide fusion inhibitors derived from 1,3,3-trimethyl-5-hydroxy-cyclohexylmethylamine is profiled. Axial disposition of the thioamide moiety is essential for potent influenza inhibitory activity.     

Plasmin activates pro-matrix metalloproteinase-2 with a membrane-type 1 matrix metalloproteinase-dependent mechanism

Membrane-type 1 matrix metalloproteinase (MT1-MMP) has been implicated as a physiological activator of progelatinase A (MMP-2). We previously reported that plasmin treatment of cells results in proMMP-2 activation and increased type IV collagen degradation. Here, we analyzed the role of MT1-MMP in plasmin activation of MMP-2 using HT-1080 cells transfected with MT1-MMP sense or antisense cDNA. Control, vector-transfected cells that expressed endogenous MT1-MMP, and antisense cDNA transfectants with very low levels of MT1-MMP did not activate proMMP-2. Conversely, cells transfected with sense MT1-MMP cDNA expressed high MT1-MMP levels and processed proMMP-2 to 68/66-kDa intermediate activation products. Control cells and MT1-MMP transfectants had much higher levels of cell-associated MMP-2 than antisense cDNA transfectants. Addition of plasmin(ogen) to control or MT1-MMP-transfected cells generated active, 62-kDa MMP-2, but was ineffective with antisense cDNA transfectants. The effect of plasmin(ogen) was prevented by inhibitors of plasmin, but not by metalloproteinase inhibitors, implicating plasmin as a mechanism for proMMP-2 activation independent of the activity of MT1-MMP or other MMPs. Plasmin-mediated activation of proMMP-2 did not result from processing of proMT1-MMP and did not correlate with alpha(v)beta(3) integrin or TIMP-2 levels. Thus, plasmin can activate proMMP-2 only in the presence of MT1-MMP; however, this process does not require the catalytic activity of MT1-MMP.     

Flora Palinologica Italiana,Sezione Aeropalinologica—S 303:Carpinus betulus L

Riassunto Nell'ambito del progetto «Flora Palinologica Italiana» e proseguendo lo studio delle Corylaceae, è qui presentata la scheda diCarpinus betulus L., con alcune osservazioni sul numero/posizione delle aperture e sulle principali differenze tra polline «fresco» e polline acetolizzato.

---

Summary According to the program «Palynological Italian Flora», and continuing the study of the Italian Corylaceae, the palynological card ofCarpinus betulus L. is shown. The study is carried out on pollen coming from three different localities and regards both «fresh» and acetolyzed pollen. For each sample, measurements are carried out on 30 «fresh» pollen grains in glicerol jelly and on 30 acetolyzed pollen grains in water/glicerol 1/1; general observations regard 500 «fresh» and 500 acetolyzed pollen grain/sample. Some morphological observations on the aperture number/position and differencies between «fresh» and acetolized pollen are pointed out.

     

Screening of Saccharomyces cerevisiae strains with respect to anaerobic growth in non-detoxified lignocellulose hydrolysate

A microplate screening method was used to assess anaerobic growth of 12 Saccharomyces cerevisiae strains in barley straw, spruce and wheat straw hydrolysate. The assay demonstrated significant differences in inhibitor tolerance among the strains. In addition, growth inhibition by the three hydrolysates differed so that wheat hydrolysate supported growth up to 70%, while barley hydrolysate only supported growth up to 50%, with dilute-acid spruce hydrolysate taking an intermediate position.     

Distinct footprints of TCR engagement with highly homologous ligands

T cell receptor engagement promotes proliferation, differentiation, survival, or death of T lymphocytes. The affinity/avidity of the TCR ligand and the maturational stage of the T cell are thought to be principal determinants of the outcome of TCR engagement. We demonstrate in this study that the same mouse TCR preferentially uses distinct residues of homologous peptides presented by the MHC molecules to promote specific cellular responses. The preference for distinct TCR contacts depends on neither the affinity/avidity of TCR engagement (except in the most extreme ranges), nor the maturity of engaged T cells. Thus, different portions of the TCR ligand appear capable of biasing T cells toward specific biological responses. These findings explain differences in functional versatility of TCR ligands, as well as anomalies in the relationship between affinity/avidity of the TCR for the peptide/MHC and cellular responses of T cells.     

Inbreeding depression in perennial Lychnis viscaria (Caryophyllaceae): effects of population mating history and nutrient availability

We studied inbreeding depression in a perennial plant, Lychnis viscaria, in three populations differing in their inbreeding history and population size by measuring several traits at two nutrient levels over the plant's life cycle. The observed levels of inbreeding depression (cumulative inbreeding depression, from -0.057 to 0.629) were high for a plant with a mixed mating system. As expected, the population with a low level of isozyme variation expressed the least inbreeding depression for seed germination. Highest inbreeding depression for germination was found in the largest and genetically most variable population. No clear differences between populations in expression of inbreeding depression in the later life stages were found. The population level inbreeding depression varied with the nutrient conditions and among populations and life stages, but we found no evidence that inbreeding depression increased with lower nutrient availability. These results emphasize the importance of measuring inbreeding depression under several environmental conditions and over life stages.     

Effect of substitution pattern on 1H, 13C NMR chemical shifts and 1J(CH) coupling constants in heparin derivatives

1H, 13C NMR chemical shifts and 1J(CH) coupling constants were measured for derivatives of heparin containing various sulfation patterns. 1H and 13C chemical shifts varied considerably after introducing electronegative sulfate groups. Chemical shifts of protons linked to carbons changed by up to 1 ppm on substitution with O- and N-sulfate or acetyl groups. Differences up to 10 ppm were detected for 13C chemical shifts in substituted glucosamine, but a less clear dependence was found in iduronate. 1J(CH) values formed two groups, corresponding to either sulfation or non-sulfation at positions 2 and 3 of glucosamine. O-sulfation caused increases up to 6 Hz in 1J(CH) and N-sulfation decreases up to 4 Hz. N-acetylation gave similar 1J(CH) values to N-sulfation. At positions 2 and 3 of iduronate the trend was less marked; 1J(CH) for O-sulfated positions usually increasing. Introduction of sulfate groups influences chemical shift and 1J(CH) values at the position of substitution, but also at more remote positions. 1J(CH) at the glycosidic linkage positions varied between free-amino and N-sulfated compounds, by up to 9 Hz. These results and changes in chemical shift values suggest that iduronate residues and the glycosidic linkages are affected, indicating overall conformational change. This may have important implications for biological activities.     

Effect of nutrient enrichment on bacterioplankton biomass and community composition in mesocosms in the Archipelago Sea, northern Baltic

Effects of nutrient enrichment on the biomass and communitycomposition of heterotrophic bacteria and picocyanobacteriawere studied in large (42 m³) mesocosms in the brackish-waterArchipelago Sea (Baltic Sea) in late summer 2000 using cellcounts and denaturing gradient gel electrophoresis (DGGE) ofpolymerase chain reaction (PCR)-amplified 16S rRNA gene fragments.The identity of the major DNA bands was determined by sequencing.The obtained sequences were related to - and -proteobacteria,actinobacteria, verrucomicrobia and cyanobacteria. Nitrogenand phosphorus additions increased the biomasses of heterotrophicbacteria and picocyanobacteria and caused significant changesin their community composition judging from the DGGE bandingpatterns. Most verrucomicrobial bands had their highest relativeintensity in the control treatment and their lowest in the highernutrient addition treatment, whereas most Synechococcus-relatedbands had their lowest relative intensity in the lower nutrientaddition treatment. The responses of proteobacteria and actinobacteriawere more variable. The presence of both freshwater and marinesequences among the closest relatives to our sequences highlightsthe intermediate character of the Archipelago Sea between afreshwater and truly marine environment.